Compositions comprising cyclosporin

ABSTRACT

The present invention relates to a formulation comprising a pharmaceutically active ingredient and coating. The invention also relates to the use of the formulation in the treatment and prevention of disorders of the gastrointestinal tract. Also disclosed are methods for preparing the formulations.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. application Ser. No.15/525,031, filed May 5, 2017, which is the U.S. National Stage ofInternational Application No. PCT/EP2015/075984, filed Nov. 6, 2015,which was published in English under PCT Article 21(2), which in turnclaims the benefit of GB Application No. 1419849.3, filed Nov. 7, 2014,International Application No. PCT/EP2014/074054, filed Nov. 7, 2014, andGB Application No. 1507673.0, filed May 5, 2015.

This invention relates to a composition comprising a pharmaceuticallyactive ingredient and a surfactant. The invention also relates to theuse of the composition in the treatment and prevention of disorders, forexample disorders of the gastrointestinal tract. Also disclosed aremethods for preparing the compositions.

BACKGROUND

Cyclosporin A is a cyclic polypeptide which has immunosuppressive andanti-inflammatory properties. The compound has been approved for theprevention of organ rejection following kidney, liver, heart, combinedheart-lung, lung or pancreas transplantation, for the prevention ofrejection following bone marrow transplantation; the treatment andprophylaxis of Graft Versus Host Disease (GVHD); psoriasis; atopicdermatitis, rheumatoid arthritis and nephrotic syndrome (Neoral™ Summaryof Product Characteristics 24 Feb. 2012). Cyclosporin A may also beuseful for the treatment of a range of other diseases including for thetreatment of severe recalcitrant plaque psoriasis Bechet's disease,anemia, myasthenia gravis and various conditions affecting the GI tract,including irritable bowel syndrome, Crohn's disease, colitis, includingulcerative colitis, diverticulitis, pouchitis, proctitis,Gastro-Intestinal Graft Versus Host Disease (GI-GVHD), colorectalcarcinoma and adenocarcinoma as well as ischemia induced disease. Arange of other diseases may benefit from treatment with cyclosporin A(Landford et al. (1998) Ann Intern Med; 128: 1021-1028) the entirety ofwhich is incorporated herein by reference. Cyclosporin A has been usedto treat a number of gastrointestinal conditions including inflammatorybowel disease (Sandborn W J, a critical review of cyclosporin therapy ininflammatory bowel disease, Inflamm Bowel Dis. 1995; 1:48-63), includingulcerative colitis (Lichtiger et al, preliminary report (cyclosporine inthe treatment of severe ulcerative colitis), Lancet. 1990; 336:16-19;Cohen et al, Intravenous cyclosporine in ulcerative colitis (a five-yearexperience), Am J Gastroenterol. 1999; 94:1587-1592).

However cyclosporin A has a number of undesirable side effects includinghypertension, impaired renal function, and neurotoxicity (Feutren et al,Risk factors for cyclosporine-induced nephropathy in patients withauto-immune diseases, International kidney biopsy registry ofcyclosporine for autoimmune diseases, N Engl J Med. 1992; 326:1654-1660;Wijdicks et al., Neurotoxicity in liver transplant recipients withcyclosporine immunosuppression, Neurology. 1995; 45:1962-1964; andPorter et al, Cyclosporine-associated hypertension, National High BloodPressure Education Program. Arch Intern Med. 1990; 150:280-283).

Cyclosporin A is available as an intravenous formulation; Sandimmun™,which is a solution of 50 mg/ml of cyclosporin A in ethanol andpolyethoxylated castor oil (for example Kolliphor™ EL). The product isalso available as orally administered formulations, including a softgelatin capsule containing a solution of cyclosporin A in ethanol, cornoil and lineoyl macrogolglycerides (Sandimmune™ Soft Gelatin capsules)and as an orally administered solution containing the cyclosporindissolved in olive oil, ethanol, and labrafil M 1944 CS (polyethoxylatedoleic glycerides) (Sandimmune™ Oral Solution). More recently amicroemulsion concentrate formulation has been approved containingcyclosporin A dissolved in DL-α-tocopherol, absolute ethanol, propyleneglycol, corn oil-mono-di-triglycerides, polyoxyl 40 hydrogenated castoroil (Neoral™). Following oral administration the Neoral™ formulationresults in the formation of a microemulsion and is stated to have animproved bioavailability compared to orally administered Sandimmune™.These orally administered cyclosporin A compositions are all instantrelease compositions and cyclosporin A will be present at highconcentration in the stomach and small intestine from where it issystemically absorbed.

Sandborn et al. (J Clin Pharmacol. 1991; 31:76-80) determined therelative systemic absorption of cyclosporin following oral andintravenous as well as oil- and water-based enemas. Based on negligibleplasma cyclosporin concentrations observed following enemaadministration, it was suggested that cyclosporin, even whensolubilised, is poorly absorbed from the colon. The enemas howeverdemonstrated considerable efficacy in the treatment of inflammatorybowel disease (Ranzi T, et al, Lancet 1989; 2:97). Intravenous or orallyadministered cyclosporin efficacy in the treatment of inflammatory boweldisease is dose dependent, requiring high doses to ensure adequateconcentration reaches the colon. Systemic toxicity is known to be doseand duration dependent.

Formulating pharmaceutically active ingredients into a form suitable foradministration to a patient is a developed area of science. It is also akey consideration for the efficacy of a drug. There are many examples ofmethods for formulating drugs and other active ingredients. The aim ofthese formulations are varied and can range from increasing systemicabsorption, allowing for a new route of administration, improvingbioavailability, reducing metabolism of the active, or avoidingundesirable routes of administration.

WO 2008/122965 discloses oral cyclosporin minicapsule compositions withmodified release properties which release cyclosporin in at least thecolon. WO2010/133609 discloses compositions comprising a water-solublepolymer matrix in which are dispersed droplets of oil, the compositionscomprising a modified release coating. The disclosed compositions alsocontain an active principle.

There remains a need for orally administered cyclosporin A compositionswhich provide high levels of cyclosporin A in the lower GI tract,particularly in the colon and absorption of the cyclosporin A from theluminal contents into the tissues of the GI tract, particularly into thecolonic tissue, for the treatment of conditions of the lower GI tractsuch as ulcerative colitis. Such compositions desirably minimise thesystemic blood exposure to cyclosporin A thereby minimising theundesirable side effect associated with systemic exposure to cyclosporinA. Particularly there is a need for orally administered compositionswhich have a low exposure/area under the curve (AUC) and/or low peakblood concentration (Cmax) compared to the orally administered productNeoral™ and/or cyclosporin A administered intravenously as for exampleSandimmune™.

Cyclosporin A is oil soluble; it is hydrophobic. Upon contact of acyclosporin solution with water, the cyclosporin can form a solid byprecipitating or crystalising out of solution. Precipitation orcrystallisation of cyclosporin from solution can occur when thecyclosporin solution is present in an oil-in-water emulsion. Prior artoil-in-water emulsions comprising cyclosporin in solution have beenfound to have low emulsion stability and to precipitate or crystalisecyclosporin over time. Solid cyclosporin formation in an emulsion isundesirable. Therefore, there is a need for emulsions comprisingcyclosporin in solution which have a higher emulsion stability. Forexample, there is a need for emulsions where the length of time forcrystal formation or precipitation to occur is longer.

Formulating an active ingredient into a bead by passing a compositioncomprising a water-soluble polymer matrix in which are disperseddroplets of oil through a single orifice nozzle is disclosed inWO2010/133609. Solid cyclosporin formation in an emulsion isparticularly undesirable whilst forming beads from an emulsioncomprising solid cyclosporin. This is because solid cyclosporin is lessactive at an intended therapeutic site, for example the gastrointestinaltract, than solubilised cyclosporin. The problem of cyclosporincrystallising or precipitating is particularly relevant in scale upproduction of beads from the emulsion. Scaling up production of beadsresults in batches of the emulsion being kept for longer periods and thepropensity for crystallisation or precipitation to happen increasing.Therefore, there is a need for an emulsion with a high stability toreduce the amount of solid in the emulsion whilst being processed intobeads and consequently to reduce the amount of solid in the bead.

Similarly there is a need for a component of the emulsion to inhibitcrystallisation or precipitation.

BRIEF SUMMARY OF THE DISCLOSURE

It has surprisingly been found that a surfactant has a stabilisingeffect on an emulsion comprising cyclosporin in solution. The surfactantmay comprise or may be a medium chain or long chain fatty acid mono- ordi-glyceride or a combination thereof, optionally wherein the surfactantdoes not comprise or is not a polyethyleneglycol ether or ester. Thesurfactant may have an HLB of up to 10, preferably of up to 7 or up to5. Specifically, it has been found that a medium chain or long chainfatty acid mono- or di-glyceride or a combination thereof, optionallynot comprising or being a polyethyleneglycol ether or ester, has astabilising effect on an emulsion comprising cyclosporin in solution.The emulsion may be an oil-in-water emulsion.

A medium chain or long chain fatty acid mono- or di-glyceride is anester of such a fatty acid and glycerol, wherein there may be one(mono-) or two (di-) fatty acids esterified to glycerol. The glycerolmay consist of one glycerol for example mono glycerol (a mono-glycerolester is also be referred to as a glyceride).

It has been found that the use of certain surfactants during themanufacture of the compositions are particularly effective instabilising the colloid (for example emulsion), resulting from themixing of an aqueous solution comprising a hydrogel forming polymer andan oil phase comprising cyclosporin A. When the colloid comprises anoil-in-water emulsion, it has been found that the presence of asurfactant having an HLB of up to 8 (particularly up to 6 or from 2 to6) in the oil phase is particularly effective in stabilising theemulsion in particular during the preparation of the composition. Thepresence of such surfactants has been found to inhibit the formation ofcyclosporin A crystals after the formation of the colloid (oil-in-wateremulsion). The presence of a surfactant with an HLB of up to 10maintains the cyclosporin A in solution in the oil phase duringmanufacture and may also provide favourable release of the cyclosporin Ain a solubilised form from the composition following oral administrationof the composition to a subject. Compositions comprising a surfactant ofthe invention with an HLB of up to 8 in at least the oil phase mayexhibit high rates of release and/or extent of release of cyclosporin Afrom the composition compared to the use of surfactants with a higherHLB value in the oil phase. The presence of a surfactant with an HLB ofup to 8 in at least the oil phase in the composition may inhibit theprecipitation of cyclosporin A after release of the cyclosporin from thecomposition thereby retaining higher levels of cyclosporin in asolubilised form within the GI tract, for example in the colon. Thecompositions described herein wherein the composition comprises an oilphase and a surfactant having an HLB of up to 10 form a furtherindependent aspect of the invention.

Accordingly, there is provided a use of a surfactant for stabilising anemulsion. Preferably, the emulsion is an oil in water emulsion. Thesurfactant preferably is or comprises a medium chain or long chain fattyacid mono- or di-glyceride or a combination thereof, optionally notcomprising or being a polyethyleneglycol ether or ester.

By the phrase “the surfactant stabilises the emulsion”, it is meant thatthe surfactant increases the length of time for solid particles (forexample precipitation or crystallisation) to occur in an emulsioncomprising cyclosporin in solution.

It has also been found that a surfactant inhibits crystallisation ofcyclosporin from a cyclosporin solution in an oil phase in anoil-in-water emulsion. The surfactant is or comprises a medium chain orlong chain fatty acid mono- or di-glyceride or a combination thereof,optionally wherein the surfactant does not comprise or is not apolyethyleneglycol ether or ester. Accordingly, the use of thesurfactant provides an emulsion that is free of crystals or precipitefor a longer period than prior art emulsions. An emulsion that takeslonger for crystallisation or precipitation to occur is beneficial forlarge-scale bead production. Consequently, the invention contemplates aprocess utilising an emulsion comprising the surfactant to producebeads, particularly in scale up of the bead production process.

Ordinarily surfactants with a low HLB value, for example up to 8, areused as water-in-oil emulsifiers. As part of the invention it has beenfound that a surfactant is an emulsifier for oil-in-water emulsionscomprising dissolved cyclosporin, wherein the surfactant has a HLB valueof up to 8 and is or comprises a medium chain or long chain fatty acidmono- or di-glyceride or a combination thereof, optionally notcomprising or being a polyethyleneglycol ether or ester.

In an aspect of the invention there is provided a liquid compositioncomprising an aqueous phase, a surfactant and an oil phase in whichcyclosporin is dissolved. The surfactant may comprise or be a mediumchain or long chain fatty acid mono- or di-glyceride or a combinationthereof and does not comprise or is not a polyethyleneglycol ether orester. The aqueous phase may comprise a hydrogel forming polymer. Theoil phase may be dispersed in the aqueous phase. The oil phase may bedispersed in the aqueous phase in the form of a colloid, for example aliquid-liquid colloid. The oil phase may be dispersed in the aqueousphase in the form of an emulsion. Accordingly, the liquid compositionmay be a liquid emulsion composition.

The liquid composition may be converted into a solid form by allowingthe hydrogel forming polymer to form a hydrogel matrix. There is alsoprovided a process for converting the liquid composition into a bead,wherein the liquid composition is ejected through a single orificenozzle.

In an aspect of the invention there is provided a composition comprisingcyclosporin, a hydrogel forming polymer matrix, a surfactant and an oilphase being dispersed in the hydrogel forming polymer matrix. Thesurfactant may be or may comprise a medium chain or long chain fattyacid mono- or di-glyceride or a combination thereof and may not compriseor may not be a polyethyleneglycol ether or ester. The composition maybe a solid composition. The composition may be in the form of a driedbead. The composition may be in the form of a dried colloid.

Advantageously an enhanced release profile is provided by the presenceof the surfactant being or comprising a medium chain or long chain fattyacid mono- or di-glyceride or a combination thereof that does notcomprise or is not a polyethyleneglycol ether or ester in compositionsof the invention compared to compositions with a different surfactant. Asolid composition of the invention exhibits a release profile withhigher release of cyclosporin and maintenance of high cyclosporin levelsin solution when compared to compositions with a different surfactant(Kolliphor EL, a polyethoxylated castor oil). The dissolution may bemeasured in deionised water.

Optionally, the cyclosporin, the hydrogel forming polymer matrix, thesurfactant and the oil phase are comprised within a core. Thus, thecomposition may comprise a core. Accordingly, the composition maycomprise a core, wherein the core comprises cyclosporin, a hydrogelforming polymer matrix, a surfactant and an oil phase being dispersed inthe hydrogel forming polymer matrix, wherein the surfactant may be orcomprise a medium chain or long chain fatty acid mono- or di-glycerideor a combination thereof and does not comprise or is not apolyethyleneglycol ether or ester.

The liquid composition may be a colloid, i.e. it may be a colloidalliquid composition. The composition may be a solid colloid or thecomposition may be in the form of a solid colloid, i.e. it may be asolid colloidal composition. The colloidal liquid composition of theinvention may comprise a continuous phase which is or comprises ahydrogel-forming polymer and a disperse phase which is or comprisescyclosporin A and an oil phase, wherein the colloidal liquid compositionor the solid colloidal composition further comprise a surfactant (alsoreferred to as a first surfactant) comprising or being a medium chain orlong chain fatty acid mono- or di-glyceride or a combination thereof andnot comprising or not being a polyethyleneglycol ether or ester.

The solid colloidal composition of the invention may comprise acontinuous phase which is or comprises a hydrogel-forming polymer matrixand a disperse phase which is or comprises cyclosporin A and an oilphase, wherein the colloidal liquid composition or the solid colloidalcomposition further comprise a surfactant (also referred to as a firstsurfactant) comprising or being a medium chain or long chain fatty acidmono- or di-glyceride or a combination thereof and not comprising or notbeing a polyethyleneglycol ether or ester.

Throughout the specification both the liquid composition and thecomposition are referred to by “composition”. Furthermore, where anembodiment or aspect is referred to as a “composition” this mayoptionally be referring to a liquid composition (for example a colloidalliquid composition) and/or to a solid composition (for example a solidcolloidal composition).

In an embodiment the oil phase comprises a solution of the cyclosporin.As such, the cyclosporin may be dissolved in the oil phase, for examplecompletely dissolved, substantially completely dissolved, or partiallydissolved. Thus, the oil phase may comprise a solution of cyclosporinand some undissolved cyclosporin.

Throughout this specification the term cyclosporin may be referring tothe class of compounds or to cyclosporin A. Preferably, the use ofcyclosporin is in reference to cyclosporin A.

The cyclosporin is suitably present in the composition in an amount offrom about 5% to about 20%, from about 8% to about 15%, or from about 9%to about 14% by weight based upon the dry weight of the core or of thecomposition.

The cyclosporin is suitably present in the liquid composition in anamount of up to 10%, optionally from about 1% to about 10%, from about2% to about 8%, from about 3% to about 6%, from about 3% to about 5% byweight of the liquid composition. Optionally the cyclosporin may bepresent in the liquid composition in about 4% by weight of the liquidcomposition.

A medium chain fatty acid mono-ester or di-ester comprises a fatty acidhaving 8 to 12 in chain carbon atoms. A long chain fatty acid mono-esteror di-ester comprises a fatty acid having at least 13 in chain carbonatoms, preferably 13 to 26 in chain carbon atoms. The long chain fattyacid may optionally have from 14 to 22 in chain carbon atoms or 16 to 20in chain carbon atoms.

Preferably, the surfactant is a medium chain or long chain fatty acidmono- or di-glyceride or a combination thereof that does not comprise oris not a polyethyleneglycol ether or ester. Where the surfactantcomprises a medium chain or long chain fatty acid mono- or di-glycerideor a combination thereof that does not comprise or is not apolyethyleneglycol ether or ester, the medium chain or long chain fattyacid mono- or di-glyceride or a combination thereof is substantially allof the surfactant. For example, the surfactant may comprise medium chainor long chain fatty acid mono- or di-glyceride or a combination thereofthat does not comprise or is not a polyethyleneglycol ether or ester inan amount of greater than 80% of the surfactant, optionally greater than85%, 90%, 95%, 97%, 98% or 99%. Suitably, the surfactant issubstantially free of a triglyceride. For example, the surfactant maycomprise less than 10%, 8%, 5%, 3%, 2% or 1% of a triglyceride.

The presence of the surfactant may enhance the rate and or extent ofrelease of cyclosporin from the composition following oraladministration. The presence of the surfactant may act to maintain ahigh proportion of the cyclosporin in a solubilised form after it hasbeen released from the composition into an aqueous medium such as thatfound in the lower GI tract, particularly the colon.

The surfactant may have an HLB value of up to 8, up to 6, or up to 5.Alternatively the surfactant may have an HLB value selected from: up to7, 1-8, 1-7, 2-6, 1-5, 2-5, 1-4, 1-3, 1-2, 2-4, 3-4, 3-6, 5-8, 6-8 and6-7. Preferably, the surfactant has an HLB value of up to 6, 2-6 or 3-6.

The cyclosporin A may be soluble in the surfactant, for example thecyclosporin A may have a solubility of more than 200 mg/g in thesurfactant. Thus, the surfactant may have a cyclosporin solubility ofmore than 200 mg/g. The surfactant may have a cyclosporin solubility offrom 200 mg/g to 500 mg/g, optionally from 250 mg/g to 500 mg/g.

The surfactant may have a cyclosporin solubility of from 200 mg/g to 400mg/g, from 225 mg/g to 375 mg/g, from 200 mg/g to 300 mg/g, from 300mg/g to 400 mg/g, from 225 mg/g to 275 mg/g, from 350 mg/g to 400 mg/g.Preferably, the surfactant has a cyclosporin solubility of from 200 mg/gto 400 mg/g or from 225 mg/g to 375 mg/g. The surfactant may have acyclosporin solubility of from 250 mg/g to 400 mg/g, from 250 mg/g to375 mg/g, from 250 mg/g to 300 mg/g, from 300 mg/g to 400 mg/g, from 250mg/g to 275 mg/g, from 350 mg/g to 400 mg/g. Preferably, the surfactanthas a cyclosporin solubility of from 250 mg/g to 400 mg/g or from 250mg/g to 375 mg/g. The solubility of cyclosporin in a surfactant may bedetermined by techniques known to those skilled in the art, for exampleby following the protocol described in Development of a SelfMicro-Emulsifying Tablet of Cyclosporine-A by the Liquisolid CompactTechnique, Zhao et al (International Journal of Pharmaceutical Sciencesand Research, 2011, Vol. 2(9), 2299-2308) which is incorporated hereinby reference.

The surfactant may have an HLB of up to 6 and a cyclosporin solubilityof from 200 mg/g to 400 mg/g. The surfactant may have an HLB value of2-6 (optionally 3-6) and a cyclosporin solubility of from 200 mg/g to400 mg/g. The surfactant may have an HLB value of 2-6 (optionally 3-6)and a cyclosporin solubility of from 225 mg/g to 275 mg/g. Thesurfactant may have an HLB value of 2-6 (optionally 3-6) and acyclosporin solubility of from 250 mg/g to 300 mg/g.

The surfactant may have an HLB of up to 6 and a cyclosporin solubilityof from 250 mg/g to 400 mg/g. The surfactant may have an HLB value of2-6 (optionally 3-6) and a cyclosporin solubility of from 250 mg/g to400 mg/g. The surfactant may have an HLB value of 2-6 (optionally 3-6)and a cyclosporin solubility of from 250 mg/g to 375 mg/g. Thesurfactant may have an HLB value of 2-6 (optionally 3-6) and acyclosporin solubility of from 250 mg/g to 300 mg/g.

The surfactant may be or comprise a medium chain or long chain fattyacid mono- or di-glyceride or a combination thereof and may not compriseor may not be a polyethyleneglycol ether or ester, wherein the fattyacid ester is saturated or unsaturated. Preferably, the fatty acid isunsaturated. The unsaturated fatty acid may contain only one or only twodouble bonds.

Where the surfactant is a medium chain or long chain fatty aciddi-glyceride (by which it is meant that there are two fatty acidsesterified to a glycerol) the surfactant may comprise two fatty acidswhich are the same or different. For example the two fatty acids mayboth be unsaturated or may both be saturated. Alternatively, one of thetwo fatty acids may be saturated and the other fatty acid may beunsaturated.

Preferably the surfactant is a long chain mono- or di-glyceride or acombination thereof and does not comprise or is not a polyethyleneglycolether or ester. A further preferred surfactant is a long chain mono- ordi-glyceride or a combination thereof and does not comprise or is not apolyethyleneglycol ether or ester, wherein the fatty acid has a chainlength of 13 to 22 carbon atoms, optionally 16 to 20 carbon atoms. Inparticular the fatty acid may have a chain length of 18 carbon atoms.

In an embodiment the surfactant is selected from: glyceryl monocaprate,glyceryl dicaprate, glyceryl monocaprylate, glyceryl dicaprylate,glyceryl caprate, glyceryl monocaprylate/caprate, glycerylcaprylate/caprate glyceryl dicaprylate/caprate, glycerylmonooleate/dioleate, glyceryl monooleate, glyceryl dioleate, glycerylmonostearate, glyceryl distearate, glyceryl monopalmitostearate,glyceryl dipalmitostearate, glyceryl monobehenate, glyceryl dibehenate,glycerol monolinoleate, glyceryl dilinoleate, polyglyceryl dioleate,propylene glycol monoheptanoate, and a combination thereof.

A preferred surfactant may be or comprise a surfactant selected from:glyceryl monocaprylate/caprate, glyceryl dicaprylate/caprate, glycerylmonooleate, glycerol monolinoleate, glyceryl dioleate, glycerylmonostearate, glyceryl distearate, glyceryl monopalmitostearate,glyceryl dipalmitostearate, glyceryl monobehenate, glyceryl dibehenate,glyceryl monolinoleate, glyceryl dilinoleate, polyglyceryl dioleate anda combination thereof.

Accordingly, there is provided a composition comprising cyclosporin, ahydrogel forming polymer matrix, a surfactant and an oil phase beingdispersed in the hydrogel forming polymer matrix, wherein the surfactantmay be or may comprise a surfactant selected from: glycerylmonocaprylate/caprate, glyceryl dicaprylate/caprate, glycerylmonooleate, glycerol monolinoleate, glyceryl dioleate, glycerylmonostearate, glyceryl distearate, glyceryl monopalmitostearate,glyceryl dipalmitostearate, glyceryl monobehenate, glyceryl dibehenate,glyceryl monolinoleate, glyceryl dilinoleate, polyglyceryl dioleate anda combination thereof.

The surfactant may comprise or be a surfactant selected from: glycerylcaprylate, glyceryl caprate, glyceryl monooleate, glyceryl dioleate,glycerol monolinoleate or a combination thereof.

A particularly preferred surfactant may be or comprise a surfactantselected from: glyceryl caprylate/caprate (Capmul MCM), glycerylmonooleate/dioleate (Capmul GMO-50) and glycerol monolinoleate (Maisine35-1).

Optionally, the surfactant is not a mixture of glyceryl monostearateEP/NF and PEG-75 palmitostearate (for example Gelto™ 64). Suitably, thesurfactant may not be or comprise a mixture of glyceryl monostearate.

In an embodiment the oil phase comprises an oil or liquid lipid and thesurfactant is present in an amount greater than the oil or liquid lipid.Optionally, the surfactant may be present in an amount of more than 6 wt% of the dry weight of the composition. This refers to the uncoatedcomposition or the core. The surfactant may comprise more than 12 wt %of the oil phase, for example in the liquid composition. The surfactantmay be present in the composition in an amount of from about 5% to about20%, from about 8% to about 20%, from about 8% to about 15%, or fromabout 10% to about 14% by weight based upon the dry weight of the core.It is to be understood that reference to the “dry weight of the core”means the weight of the components present in the uncoated core otherthan water.

The weight ratio of the surfactant:oil may be from about 5:1 to about1:5, from about 3:1 to about 1:2, from about 3:1 to about 1:1 or fromabout 2.5:1 to 1.5:1. Suitably the weight ratio may be about 1:1, about2:1, about 2.5:1, about 3:1, about 1:1.5 or about 1:2.

Accordingly, in a preferred embodiment there is provided a liquidcomposition comprising an aqueous phase, a surfactant and an oil phasein which cyclosporin is dissolved, wherein the aqueous phase comprises ahydrogel forming polymer, the oil phase is dispersed in the aqueousphase and the surfactant comprises or is a surfactant selected from:glyceryl caprylate/caprate (Capmul MCM), glyceryl monooleate/dioleate(Capmul GMO-50), glycerol monolinoleate (Maisine 35-1) and a combinationthereof. The oil phase may be dispersed in the aqueous phase in the formof a colloid, for example a liquid-liquid colloid. The oil phase may bedispersed in the aqueous phase in the form of an emulsion. Accordingly,the liquid composition may be a liquid emulsion composition.

There is provided a composition comprising cyclosporin, a hydrogelforming polymer matrix, a surfactant and an oil phase being dispersed inthe hydrogel forming polymer matrix, wherein the surfactant comprises oris a surfactant selected from: glyceryl caprylate/caprate (Capmul MCM),glyceryl monooleate/dioleate (Capmul GMO-50), glycerol monolinoleate(Maisine 35-1) and a combination thereof. The composition may be a solidcomposition. The composition may be in the form of a dried bead. Thecomposition may be in the form of a dried colloid.

Optionally, the surfactant is or comprises glyceryl monooleate, glyceryldioleate or a combination thereof. Capmul GMO-50 is an example of acommercially available surfactant that comprises a combination ofglyceryl monooleate and glyceryl dioleate. Thus, the surfactant may beCapmul GMO-50. Where Capmul GMO-50 is mentioned in the specification itwill be understood that it is referring to a mixture of glycerylmonooleate and glyceryl dioleate. Capmul GMO-50 may also refer toglyceryl monooleate alone.

Similarly, the skilled person would understand that a surfactant that isdescribed as, for example glyceryl monooleate/dioleate, contemplates acombination of glyceryl monooleate and glyceryl dioleate. In other wordsa “I” in a surfactant name indicates that the surfactant is a mixture oftwo components.

The composition may comprise a coating to control or modulate release ofthe cyclosporin from the composition. Advantageously the coating is apolymeric coating to provide delayed and/or sustained release of thecyclosporin from the composition. Suitably such coatings are describedin more detail below and include a coating which is or comprises acoating selected from a controlled release polymer, a sustained releasepolymer, an enteric polymer, a pH independent polymer, a pH dependentpolymer and a polymer specifically susceptible to degradation bybacterial enzymes in the gastrointestinal tract, or a combination of twoor more such polymers. In a particular embodiment the coating is orcomprises a pH-independent polymer, for example a coating which is orcomprises ethyl cellulose. In a further specific embodiment the coatingis or comprises a pH-independent polymer, for example ethyl cellulose,and optionally a water-soluble polysaccharide, for example pectin orchitosan, or a combination thereof, particularly pectin.

In an embodiment the coating that is referred to in the precedingparagraph is an outer coating, also referred to as a second coating. Thecomposition may optionally comprise a further coating, referred to as asub-coat or a first coating. The respective polymers of the firstcoating and the second coating are different. Often the second coatingdoes not have any polymer found in the first coating; for example, ifthe first coating comprises (e.g. is) a hydroxypropylmethyl cellulose,then the second coating will not also comprise a hydroxypropylmethylcellulose. In addition the situation is contemplated where the firstcoating is or comprises a water-soluble ether or ester of a celluloseether, the major component(s) (e.g. more than 50%) of the second coatingis or comprises a different polymer to that of the first coating.Accordingly, the first and second coatings suitably provide two layersof material as part of the composition. It is to be understood that whenthe second coating comprises a mixture of components, minor componentsof the outer second coating may be the same as the material of thesub-coating. By way of example, when the first coating is or comprisesHPMC and the second coating comprises ethyl cellulose, the ethylcellulose may optionally further comprise a minor amount (e.g. less than50%, 40%, 30% or 20%) of the first coating material, HPMC in thisexample. In such embodiments the first coating and the second coatingare considered to be different.

The composition of the invention may comprise cyclosporin, a hydrogelforming polymer matrix, a surfactant and an oil phase being dispersed inthe hydrogel forming polymer matrix, wherein the surfactant may be amedium chain or long chain fatty acid mono- or di-glyceride or acombination thereof and may not comprise or may not be apolyethyleneglycol ether or ester. Optionally, the composition mayfurther comprise a first coating, wherein the first coating is orcomprises a water-soluble cellulose ether as described above andelsewhere herein. In addition to the first coating or alternatively tothe first coating the composition may comprise a second coating.Optionally, the second coating is or comprises a coating, suitably apolymeric coating, to control or modulate release of the activeingredient from the composition. The polymeric coating may be as furtherdescribed elsewhere in this specification.

Where the composition comprises a first coating and a second coating thesecond coating may be outside the first coating.

The composition may comprise: a core, wherein the core comprisescyclosporin, a hydrogel forming polymer matrix, a surfactant and an oilphase being dispersed in the hydrogel forming polymer matrix; a firstcoating outside the core, wherein the first coating is a water-solublecellulose ether as described above and elsewhere herein; and a secondcoating outside the first coating, wherein the surfactant is asdescribed herein, the surfactant being, for example, a medium chain orlong chain fatty acid mono- or di-glyceride or a combination thereof andnot comprising or not being a polyethyleneglycol ether or ester.Throughout this specification “core” may refer to a core comprisingcyclosporin, a hydrogel forming polymer matrix, a surfactant, asdescribed herein, and an oil phase being dispersed in the hydrogelforming polymer matrix.

According to an embodiment of the invention, the surfactant optionallyis a medium chain or long chain fatty acid mono-glyceride, di-glycerideor a combination thereof, the first coating is or comprises awater-soluble cellulose ether, and the composition further comprises asecond coating outside the first coating wherein the second coating isor comprises a coating, suitably a polymeric coating, to control ormodulate release of the active ingredient from the composition. Thepolymeric coating may be as further described elsewhere in thisspecification.

The first coating suitably may be or comprise a water-soluble celluloseether. The water-soluble cellulose ether may be any cellulose ether orderivative of a cellulose ether, for example an ester of a celluloseether that is soluble in water. Therefore, the water-soluble celluloseether may be selected from: an alkyl cellulose; a hydroxyalkylcellulose; a hydroxyalkyl alkyl cellulose; and a carboxyalkyl cellulose.Suitably the first coating is or comprises one or more water-solublecellulose ethers selected from: methyl cellulose, hydroxyethylcellulose, hydroxypropyl cellulose and hydroxypropylmethyl cellulose,and combinations thereof. In particular embodiments the first coating isor comprises a water-soluble hydroxypropyl methylcellulose. Thewater-soluble cellulose ethers and water-soluble derivatives thereof(e.g. water-soluble esters of a cellulose ether) present in the firstcoating (sub-coat) suitably form at least 20%, 40%, 50%, 60%, 70%, 80%,85% or 90% by weight of the dry weight of the first coating.

In accordance with the present invention there is provided apharmaceutical composition comprising a core and a first coating,wherein the core comprises cyclosporin, a hydrogel forming polymermatrix, a surfactant and an oil phase being dispersed in the hydrogelforming polymer matrix and the first coating comprises or is a watersoluble cellulose ether and the first coating is present in an amountcorresponding to a weight gain due to the first coating of from 0.5% to20% by weight of the core, wherein the surfactant is as describedherein, for example a medium chain or long chain fatty acid mono- ordi-glyceride or a combination thereof and not comprising or not being apolyethyleneglycol ether or ester.

The first coating of the present invention modifies the release of theactive ingredient from the composition. There would be an expectationthat a coating on a composition would slow the rate of release of theactive ingredient within a composition. One might reasonably expect thisas coating the composition with additional material would provide anadditional barrier to a dissolution medium coming into contact with theactive ingredient in the composition. In contrast to this expectedoutcome, the compositions of the present invention comprise a coatingcomprising or being a water soluble cellulose ether that increases therate of release of the active ingredient compared to a compositionwithout the coating. In addition the coating of the present inventionhas the beneficial effect of maintaining the active ingredient insolution, whereas a comparable composition lacking the coating of theinvention provides less of the active ingredient in solution as timeprogresses. Without wishing to be bound by theory, it is believed thatthe coating prevents precipitation of the active ingredient fromsolution, thereby maintaining a higher amount of the active in solution.

Throughout the present application active ingredient, active, andpharmaceutically active ingredient are used interchangeably and allrefer to cyclosporin, preferably cyclosporin A.

The composition of the present invention may take any form known to theperson skilled in the art. Preferably, the composition is an oralcomposition. The composition may be in the form of a single minibead ora multiplicity of minibeads. Accordingly the invention provides aminibead comprising cyclosporin, a hydrogel forming polymer matrix, asurfactant and an oil phase being dispersed in the hydrogel formingpolymer matrix, wherein the surfactant is or comprises a medium chain orlong chain fatty acid mono- or di-glyceride or a combination thereof anddoes not comprise or is not a polyethyleneglycol ether or ester. Theinvention also provides a composition comprising a multiplicity ofminibeads. Similarly, the invention provides a multiple minibeadformulation comprising a unit dosage form comprising a multiplicity ofminibeads.

The invention also provides for a pharmaceutical composition comprisinga core and a first coating, wherein the core comprises cyclosporin, ahydrogel forming polymer matrix, a surfactant and an oil phase beingdispersed in the hydrogel forming polymer matrix and the first coatingcomprises or is a water-soluble cellulose ether and the first coatinghas a thickness of from 1 μm to 1 mm wherein the surfactant is asdescribed herein, for example a medium chain or long chain fatty acidmono- or di-glyceride or a combination thereof and not comprising or notbeing a polyethyleneglycol ether or ester.

Any of the pharmaceutical compositions of the invention may comprise afurther coating, referred to herein as a second coating. The secondcoating may be outside the first coating. The second coating may be orcomprise a delayed release polymer. In any embodiment and any aspect ofthe invention the first and second coating may be different.

The invention therefore, contemplates a pharmaceutical compositioncomprising a core, a first coating and a second coating outside of thefirst coating, wherein the core comprises cyclosporin, a hydrogelforming polymer matrix, a surfactant and an oil phase being dispersed inthe hydrogel forming polymer matrix, the first coating comprises or is awater soluble cellulose ether (for example HPMC), and the second coatingcomprises or is a delayed release polymer (for example ethylcellulose),wherein the surfactant is a medium chain or long chain fatty acid mono-or di-glyceride or a combination thereof and does not comprise or is nota polyethyleneglycol ether or ester.

The composition of any aspect or embodiment of the invention may be inthe form of a solid colloid. Furthermore, the core of a composition maybe in the form of a solid colloid. The colloid comprises a continuousphase and a disperse phase. Suitable continuous phases and dispersephases which may be used to form the core are defined in more detailbelow and in the detailed description of the invention. The continuousphase may comprise or be the hydrogel forming polymer matrix. Hence,where the continuous phase is the hydrogel forming polymer matrix, thecomposition of the invention may take the form of a solid unit of thehydrogel forming polymer comprising a disperse phase. The disperse phasemay be droplets dispersed in the continuous phase, or the hydrogelforming polymer matrix. The disperse phase may comprise or be the oilphase.

Thus, the invention provides a composition in the form of a solidcolloid comprising a continuous phase and a dispersed phase, wherein thecontinuous phase comprises or is a hydrogel forming polymer matrix andthe continuous phase is or comprises oil phase, wherein the compositionfurther comprises cyclosporin and a surfactant. Preferably, thesurfactant is a medium chain or long chain fatty acid mono- ordi-glyceride or a combination thereof and does not comprise or is not apolyethyleneglycol ether or ester. The oil phase may comprise thecyclosporin in solution.

The composition may comprise a core in the form of a solid colloidcomprising a continuous phase and a dispersed phase, wherein thecontinuous phase comprises or is a hydrogel forming polymer matrix andthe continuous phase is or comprises oil phase, wherein the core furthercomprises cyclosporin and a surfactant, wherein the surfactant is amedium chain or long chain fatty acid mono- or di-glyceride or acombination thereof and does not comprise or is not a polyethyleneglycolether or ester. The oil phase may comprise the cyclosporin in solution.

The continuous phase of a solid colloid composition or core is orcomprises a hydrogel-forming polymer matrix. In embodiments thehydrogel-forming polymer matrix is or comprises a hydrocolloid, anon-hydrocolloid gum or chitosan. In a particular embodiment thehydrogel-forming polymer matrix is or comprises gelatin, agar, apolyethylene glycol, starch, casein, chitosan, soya bean protein,safflower protein, alginates, gellan gum, carrageenan, xanthan gum,phthalated gelatin, succinated gelatin, cellulosephthalate-acetate,oleoresin, polyvinylacetate, polymerisates of acrylic or methacrylicesters and polyvinylacetate-phthalate and any derivative of any of theforegoing; or a mixture of two or more such polymers. In a furtherembodiment the hydrogel-forming polymer matrix is or comprises ahydrocolloid selected from carrageenan, gelatin, agar and pectin, or acombination thereof optionally selected from gelatin and agar or acombination thereof. Particularly, the polymer of the hydrogel-formingpolymer matrix is or comprises gelatin. In an embodiment, thehydrogel-forming polymer does not comprise a cellulose or a cellulosederivative, e.g. does not comprise a cellulose ether.

In this aspect of the invention the composition may be in the form of asolid colloid the colloid comprising a continuous phase and a dispersephase and the cyclosporin may be in solution or suspended in thedisperse phase. For example, the cyclosporin may be in solution in thedisperse phase.

It is to be understood that the individual embodiments described abovemay be combined with one or more of the other embodiments described toprovide further embodiments of the invention.

The first coating may be in contact with the core. The second coatingmay be on the first coating. In embodiments the first coating is incontact with the core and the second coating is on the first coating.

The second coating may be or may comprise a delayed release polymer andthe delayed release polymer may be selected from an enteric polymer, apH independent polymer, a pH dependent polymer and a polymerspecifically susceptible to degradation by bacterial enzymes in thegastrointestinal tract, or a combination of two or more such polymers.Hence, the second coating may be any of the aforementioned delayedrelease polymers or any may be or possess the characteristics mentionedin relation to the delayed release polymer mentioned below.

In embodiments the delayed release polymer may be water-soluble orwater-permeable in an aqueous medium with a pH greater than 6.5. Thedelayed release polymer may be or comprise a pH-independent polymer, forexample ethyl cellulose.

In any aspect and any embodiment of the invention the water-solublecellulose ether may be selected from any one or a combination of: methylcellulose, hydroxyethyl cellulose, hydroxypropyl cellulose andhydroxypropyl methylcellulose. The water-soluble cellulose ether maypreferably be hydroxylpropyl methylcellulose (HPMC).

In embodiments the first coating may be or comprise hydroxypropyl methylcellulose and the second coating may be or comprise ethyl cellulose.

The disclosure of the weight gain of the first coating is given as a %by weight of the core. Similarly, the weight gain of the second coatingis given as a % by weight of the core, where there is no first coating(sub-coat) on the core. Where the composition comprises a first coating,the weight gain of the second coating is given as a % by weight of thecomposition that is coated by the second coating, for example the coreand the first coating.

The hydrogel forming polymer or the hydrogel forming polymer matrix maybe or comprise a hydrocolloid, a non-hydrocolloid gum or chitosan. Thehydrogel forming polymer or the hydrogel forming polymer matrix may be areversible hydrocolloid, for example a thermoreversible hydrocolloid ora thermoreversible hydrogel forming polymer. Alternatively, the hydrogelforming polymer or the hydrogel forming polymer matrix may be orcomprise an irreversible hydrocolloid. The hydrogel forming polymer orthe hydrogel forming polymer matrix may be or comprise gelatin, agar, apolyethylene glycol, starch, casein, chitosan, soya bean protein,safflower protein, alginates, gellan gum, carrageenan, xanthan gum,phthalated gelatin, succinated gelatin, cellulosephthalate-acetate,oleoresin, polyvinylacetate, polymerisates of acrylic or methacrylicesters and polyvinylacetate-phthalate and any derivative of any of theforegoing; or a mixture of one or more such a hydrogel forming polymers.The hydrogel forming polymer or the hydrogel forming polymer matrix maybe or comprise a hydrocolloid selected from carrageenan, gelatin, agarand pectin, or a combination thereof optionally selected from gelatinand agar or a combination thereof, more optionally the hydrogel formingpolymer or the or the hydrogel forming polymer matrix forming polymermatrix is or comprises gelatin. The hydrogel forming polymer matrix isor comprises a non-hydrocolloid gum optionally selected from across-linked salt of alginic acid. In preferred embodiments the hydrogelforming polymer or the hydrogel forming polymer matrix is or comprisesgelatin.

In embodiments the hydrogel forming polymer or the hydrogel formingpolymer matrix further comprising a plasticiser, optionally aplasticiser selected from glycerin, a polyol for example sorbitol,polyethylene glycol and triethyl citrate or a mixture thereof,particularly sorbitol.

The hydrogel forming polymer matrix may encapsulate the cyclosporin. Thecyclosporin may be encapsulated in solution. The cyclosporin may be insolution or suspended in another component, for example the oil phase orthe disperse phase discussed elsewhere, of the composition that is alsoencapsulated by the hydrogel forming polymer matrix.

The disperse phase may be solid, semi-solid or liquid. In particular,the disperse phase may be liquid. In other particular instances thedisperse phase may be semi-solid, for example it may be waxy.

The disperse phase may be or comprise the oil phase, for example the oilphase may be a solid, a semi-solid or a liquid. Suitably the dispersephase or the oil phase is or comprises a liquid lipid and optionally asolvent miscible therewith. The liquid lipid is optionally a mediumchain mono- di- or triglyceride (particularly a medium chaintriglyceride).

Suitably, cyclosporin is soluble in the solvent. The solvent may be analcohol (for example ethanol or isopropanol), a glycol (for examplepropylene glycol or a polyethylene glycol) or a glycol ether. Thesolvent may be a glycol ether, for example an ethylene glycol ether,more particularly an alkyl, aryl or aralkyl ethylene glycol ether. Thesolvent may be a glycol ether selected from 2-methoxyethanol;2-ethoxyethanol; 2-propoxyethanol; 2-isopropoxyethanol; 2-butoxyethanol;2-phenoxyethanol; 2-benzyloxyethanol; 2-(2-methoxyethoxy)ethanol;2-(2-ethoxyethoxy)ethanol; and 2-(2-butoxyethoxy)ethanol. Moreparticularly the solvent is 2-(2-ethoxyethoxy)ethanol or2-phenoxyethanol. A particular solvent is 2-(2-ethoxyethoxy)ethanol.

The cyclosporin may be dissolved in the disperse phase. The cyclosporinmay be suspended in the disperse phase. The disperse phase may be asdescribed elsewhere herein, for example it may be as described in theimmediately preceding two paragraphs.

The oil phase or disperse phase may be or may comprise a liquid lipid.Particularly, the oil phase or disperse phase may comprise or be ashort-, medium- or long-chain triglyceride formulation, or a combinationthereof, for example a caprylic/capric triglyceride, i.e. acaprylic/capric triglyceride formulation.

Accordingly, in an embodiment the liquid composition comprises anaqueous phase, a surfactant and an oil phase in which cyclosporin isdissolved, wherein the surfactant may comprise or be a medium chain orlong chain fatty acid mono- or di-glyceride or a combination thereof anddoes not comprise is not a polyethyleneglycol ether or ester, theaqueous phase may comprise a hydrogel forming polymer, and the oil phasecomprises a short-, medium- or long-chain triglyceride formulation, or acombination thereof (optionally a caprylic/capric triglyceride, i.e. acaprylic/capric triglyceride formulation) and is dispersed in theaqueous phase. The oil phase may be dispersed in the aqueous phase inthe form of a colloid, for example a liquid-liquid colloid. The oilphase may be dispersed in the aqueous phase in the form of an emulsion.Accordingly, the liquid composition may be a liquid emulsioncomposition.

Additionally, in an embodiment the composition comprises cyclosporin, ahydrogel forming polymer matrix, a surfactant and an oil phasecomprising a short-, medium- or long-chain triglyceride formulation, ora combination thereof (optionally a caprylic/capric triglyceride, i.e. acaprylic/capric triglyceride formulation) and being dispersed in thehydrogel forming polymer matrix, wherein the surfactant may be or maycomprise a medium chain or long chain fatty acid mono- or di-glycerideor a combination thereof and does not comprise or is not apolyethyleneglycol ether or ester. The composition may be in the form ofa dried colloid. The composition may be in the form of a bead.

In a particular embodiment the disperse phase or the oil phase furthercomprises a solvent, thus optionally the disperse phase or the oil phasemay be or comprise a liquid lipid and a solvent. The solvent may bemiscible with the liquid lipid and water, optionally wherein the solventis selected from 2-(2-ethoxyethoxy)ethanol and a poly(ethylene glycol),particularly wherein the solvent is 2-(2-ethoxyethoxy)ethanol. In afurther embodiment the disperse phase or oil phase is or comprises amedium chain mono- di- or triglyceride (particularly a medium chaintriglyceride), 2-(ethoxyethoxy)ethanol and the surfactant. The dispersephase or oil phase as described in this paragraph may contain thecyclosporin, the cyclosporin may optionally be in solution.

Preferably, the oil phase or disperse phase comprises a short-, medium-or long-chain triglyceride formulation, or a combination thereof(optionally a caprylic/capric triglyceride, i.e. a caprylic/caprictriglyceride formulation). Where the oil phase or disperse phasecomprises a short-, medium- or long-chain triglyceride formulation, or acombination thereof, the triglyceride is substantially all of thedisperse phase or oil phase (optionally the liquid lipid). For example,the oil phase or disperse phase may comprise short-, medium- orlong-chain triglyceride formulation in an amount of greater than 80% ofthe oil phase or disperse phase (optionally the liquid lipid),optionally greater than 85%, 90%, 95%, 97%, 98% or 99%. Suitably, theshort-, medium- or long-chain triglyceride formulation is substantiallyfree of mono- or di-glycerides. For example, the surfactant may compriseless than 10%, 8%, 5%, 3%, 2% or 1% of a mono- or di-glycerides.

In embodiments the composition further comprises one or more additionalsurfactants, preferably one additional surfactant. The additionalsurfactant may be referred to as a second surfactant or furthersurfactant throughout the specification and these terms are usedinterchangeably. Where the compositions of the invention comprise asecond surfactant the surfactant that may be or may comprise a mediumchain or long chain fatty acid mono- or di-glyceride or a combinationthereof and does not comprise or is not a polyethyleneglycol ether orester is referred to as a first surfactant.

Suitable surfactants for the second surfactant are described in moredetail in the detailed description of the invention. The secondsurfactant may be an anionic or non-ionic surfactant. The secondsurfactant may be a sucrose monoester, an alkyl sulfate or apolyethylene glycol alkyl ether. The second surfactant may be sucroselaurate, sucrose palmitate, sodium octyl sulfate, sodium octadecylsulfate, sodium dodecyl sulphate, polyethylene glycol hexadecyl ether,polyoxyethylene glycol octadecyl ether, or polyethylene glycol dodecylether. Optionally, the second surfactant may be sodium octyl sulfate,sodium octadecyl sulfate, sodium dodecyl sulphate or polyethylene glycoldodecyl ether.

Preferably the second surfactant is an anionic surfactant. For example,the second surfactant may be an alkyl sulphate, for example sodium octylsulfate, sodium octadecyl sulfate, or sodium dodecyl sulphate(preferably sodium dodecyl sulphate).

In those embodiments where the liquid composition is in the form of acolloid, the composition is in the form of a solid colloid or thecomposition comprises a core in the form of a solid colloid, the colloidcomprises a continuous phase and a disperse phase, wherein thecontinuous phase comprises the hydrogel-forming polymer matrix and thesecond surfactant may be present in the continuous phase, the dispersephase or both. Preferably the second surfactant is present in thecontinuous phase and the first surfactant is present in the dispersephase. Accordingly, the aqueous phase of the liquid composition maycomprise the second surfactant and the oil phase may comprise the firstsurfactant. In one embodiment the core further comprises one additionalsurfactant present in at least the continuous phase, the surfactanthaving an HLB value of greater than 10, for example greater than 20.

The composition may have the characteristics of a composition formed bymixing a disperse phase with a continuous phase to form a colloid,wherein the continuous phase is an aqueous phase comprising hydrogelforming polymer and the disperse phase is a oil phase, wherein thepharmaceutically active ingredient is in the continuous phase or thedisperse phase, wherein the colloid is gelled to form the composition.The composition is thus in the form of a solid colloid.

Furthermore, the composition may comprise a core having thecharacteristics of a core formed by mixing a disperse phase with acontinuous phase to form a colloid, wherein the continuous phase is anaqueous phase comprising hydrogel forming polymer and the disperse phaseis a oil phase, wherein the pharmaceutically active ingredient is in thecontinuous phase or the disperse phase, wherein the colloid is gelled toform the core. The core is thus in the form of a solid colloid.

The cyclosporin may be present in the composition in solution or insuspension. In the aspect where the invention provides a liquidcomposition the cyclosporin is in solution.

The liquid composition comprises an aqueous phase, a surfactant and anoil phase in which cyclosporin is dissolved and may have thecharacteristics of a liquid composition obtained by a processcomprising:

(i) dissolving a hydrogel-forming polymer in an aqueous liquid to forman aqueous phase solution;(ii) dissolving the cyclosporin in the oil phase to form a solution; and(iii) mixing the aqueous phase solution (i) and the oil phase solution(ii) to form a colloid (optionally an emulsion).

The composition or core of the invention may have the characteristics ofa composition obtained by a process comprising:

(a) ejecting the liquid composition through a nozzle to form droplets;(b) causing or allowing the a hydrogel-forming polymer to gel orsolidify to form a hydrogel-forming polymer matrix; and(c) drying the solid.

The composition or core comprises cyclosporin, a hydrogel formingpolymer matrix, a surfactant and an oil phase and may have thecharacteristics of a composition obtained by a process comprising:

(i) dissolving a hydrogel-forming polymer in an aqueous liquid to forman aqueous phase solution;(ii) dissolving the cyclosporin in the oil phase to form a solution;(iii) mixing the aqueous phase solution (i) and the oil phase solution(ii) to form a colloid (optionally an emulsion);(iv) ejecting the colloid through a nozzle to form droplets;(v) causing or allowing the a hydrogel-forming polymer to gel orsolidify to form a hydrogel-forming polymer matrix; and(vi) drying the solid.

The aqueous phase and oil phase may be mixed (for example in step (iii))in an oil phase to aqueous phase ratio of from 1:4 to 1:10, optionallyfrom 1:4 to 1:8, from 1:5 to 1:7. For example, the oil phase to aqueousphase ratio may be 1:4, 1:5, 1:6 or 1:7.

The oil phase solution (ii) may be prepared by dissolving or dispersingthe cyclosporin A in a suitable hydrophobic liquid. The hydrophobicliquid may be for example, any of the oils or liquid lipids describedherein. By way of example the hydrophobic liquid may be, or comprise,saturated or unsaturated fatty acids or a triglyceride, or an ester orether thereof with polyethylene glycols. A particular oil for the oilphase is or comprises a triglyceride, for example an oil comprising amedium chain triglyceride, optionally wherein the oil comprises atriglyceride of at least one fatty acid selected from fatty acids having6, 7, 8, 9, 10, 11 or 12 carbon atoms, e.g. 08-010 fatty acids.

The aqueous phase solution (i) may further comprise a surfactantselected from: sucrose monoester, an alkyl sulfate and a polyethyleneglycol alkyl ether, optionally the selected from: sucrose laurate,sucrose palmitate, sodium octyl sulfate, sodium octadecyl sulfate,sodium dodecyl sulphate, polyethylene glycol hexadecyl ether,polyoxyethylene glycol octadecyl ether, and polyethylene glycol dodecylether. The aqueous phase solution (i) may further comprise a surfactantselected from: sodium octyl sulfate, sodium octadecyl sulfate, sodiumdodecyl sulphate or polyethylene glycol dodecyl ether. Preferably, theaqueous phase solution (i) further comprises an anionic surfactant, e.g.as described elsewhere herein, for example sodium dodecyl sulphate(SDS).

In one embodiment the liquid composition or composition having thecharacteristics of a liquid composition or composition obtained by theprocess above is a composition or liquid composition comprising an oilphase dispersed in the aqueous phase solution, wherein the liquidcomposition or composition is or comprises cyclosporin, glycerylmonooleate/dioleate, gelatin, SDS, sorbitol, caprylic/caprictriglyceride, 2-(ethoxyethoxy)ethanol; wherein the aqueous phasesolution (i) is or comprises gelatin, sorbitol and SDS; and the oilphase solution (ii) is or comprises cyclosporin, glycerylmonooleate/dioleate, caprylic/capric triglyceride,2-(ethoxyethoxy)ethanol and the active ingredient.

Cores having the characteristics of cores obtained by theabove-described processes, for example cores obtained by the processes,may be coated to provide a coating that comprises or is a water-solublecellulose ether, optionally with a second coating to control or modifyrelease, preferably a polymeric coating as described above and herein.The coated composition may be obtained by applying to the core thecoating, e.g. applying to the core first and second coatings asdescribed. Before the coating is applied, the core may be made by aprocess having steps (i) to (vi) or (i) to (v) described above. Suitablemethods for applying the coating(s) are described below and includeapplying the coatings by spray coating a coating composition onto thecore. The processes having steps (i) to (vi) or (i) to (v) themselvesform aspects of the invention.

The composition or core may further comprise a second surfactant (alsoreferred to as a further surfactant), optionally wherein the secondsurfactant is an anionic surfactant, optionally selected from alkylsulphates, carboxylates or phospholipids, or a non-ionic surfactant,optionally selected from sorbitan-based surfactants, PEG-fatty acids,fatty alcohol ethoxylates, alkylphenol ethoxylate, fatty acidethoxylates, fatty amide ethoxylates, alkyl glucosides or glyceryl fattyacids, or poloxamers, or a combination thereof. Hence the liquidcomposition of the invention may comprise at least the followingfeatures, an aqueous phase comprising a hydrogel forming polymer, afirst surfactant and an oil phase being dispersed in the aqueous phasein which cyclosporin is dissolved and a second surfactant. Similarly,the composition of the invention may comprise at least the followingfeatures, cyclosporin, a hydrogel forming polymer matrix, a firstsurfactant and an oil phase being dispersed in the hydrogel formingpolymer matrix and a second surfactant.

In embodiments where the composition is in the form of a solid colloid,the second surfactant may be in the disperse phase or the continuousphase. The second surfactant may be in the continuous phase and may bean anionic surfactant, for example at least one surfactant selected fromfatty acid salts and bile salts, particularly an alkyl sulphate, forexample sodium dodecyl sulphate. The surfactant in the disperse phasemay be a non-ionic surfactant.

In embodiments the composition comprises a second surfactant which is orcomprises an anionic surfactant, for example sodium dodecyl sulphate,which is in the continuous phase.

In embodiments the composition further comprises a combination ofexcipients selected from: an anionic surfactant and a solvent; ananionic surfactant and an oil; and an anionic surfactant, a solvent andan oil. Preferably, the anionic surfactant is an alkyl sulphate, forexample sodium dodecyl sulphate, the oil is a medium chain mono-, di-and/or tri-glyceride (optionally a medium chain triglyceride, forexample caprylic/capric triglyceride, and the solvent is2-(ethoxyethoxy)ethanol.

The composition may further comprise an excipient selected from: asurfactant, a solubiliser, a permeability enhancer, a disintegrant, acrystallisation inhibitor, a pH modifier, a stabiliser, or a combinationthereof.

The composition of the invention or, where the composition comprises acore, the core may comprise a disperse phase or oil phase, wherein thedisperse phase or oil phase is or comprises:

cyclosporin;

a medium or long chain fatty acid mono- or di-ester or a combinationthereof which does not comprise is not a polyethyleneglycol ether orester, such as a medium or long chain fatty acid mono- or di-glycerideor a combination thereof, for example glyceryl monooleate/dioleate;

a medium chain mono- di- or tri-glyceride, for example caprylic/caprictriglyceride; and

a solvent, for example 2-(ethoxyethoxy)ethanol and the composition orthe core may further comprise a continuous phase or aqueous phase beingor comprising:

an anionic surfactant, for example at least one surfactant selected fromfatty acid salts and bile salts, particularly an alkyl sulphate, forexample sodium dodecyl sulphate

a hydrogel forming polymer matrix which is or comprises a hydrocolloidselected from carrageenan, gelatin, agar and pectin, or a combinationthereof optionally selected from gelatin and agar or a combinationthereof, more optionally the polymer of the a hydrogel forming polymermatrix is or comprises gelatin; and

optionally a plasticiser, for example a plasticiser selected fromglycerin, a polyol for example sorbitol, polyethylene glycol andtriethyl citrate or a mixture thereof, particularly sorbitol.

In one embodiment the composition comprises a core and a coating outsidethe core, wherein the core is in the form of a solid colloid, thecolloid comprising a continuous phase and a disperse phase, wherein thedisperse phase is or comprises:

cyclosporin A;

a medium or long chain fatty acid mono- or di-glyceride or a combinationthereof which does not comprise is not a polyethyleneglycol ether orester, for example glyceryl monooleate/dioleate;

a medium chain mono- di- and/or tri-glyceride, for examplecaprylic/capric triglyceride; and

a co-solvent, for example 2-(ethoxyethoxy)ethanol;

and wherein the continuous phase is or comprises:

-   -   a hydrogel-forming polymer matrix which is or comprises a        hydrocolloid selected from carrageenan, gelatin, agar and        pectin, or a combination thereof optionally selected from        gelatin and agar or a combination thereof, more optionally the        polymer of the water-soluble polymer matrix is or comprises        gelatin;    -   optionally a plasticiser, optionally a plasticiser selected from        glycerin, a polyol for example sorbitol, polyethylene glycol and        triethyl citrate or a mixture thereof, particularly sorbitol;        and

an anionic surfactant, for example at least one surfactant selected fromfatty acid salts and bile salts, particularly an alkyl sulphate, forexample sodium dodecyl sulphate;

and wherein the coating on the core is a first coating or a secondcoating, as described herein.

Suitably the coating comprises a first coating and a second coatingoutside the first coating; and wherein

the first coating is the coating which is or comprises a water-solublecellulose ether as described above; and

the second coating is or comprises a coating, suitably a polymericcoating, as defined above to control or modulate release of cyclosporinA from the composition.

In embodiments comprising a first coating and/or a second coating, forexample as mentioned in the immediately preceding paragraph, aparticular first coating is or comprises hydroxypropylmethyl celluloseand a particular second coating outside the first coating is orcomprises a pH independent polymer, for example ethyl cellulose; moreparticularly the second coating is or comprises ethyl cellulose andoptionally a polysaccharide selected from water soluble and naturallyoccurring polysaccharides, for example pectin or another water-solublenaturally occurring polysaccharide. The second coating may thereforecontain pectin or another said polysaccharide or it may be substantiallyfree of pectin and other said polysaccharides. There are thereforedisclosed second coatings which comprise ethylcellulose as a controlledrelease polymer and which further comprise pectin or another saidpolysaccharide as well as second coatings which comprise ethylcelluloseas a controlled release polymer and which do not further comprise pectinor another said polysaccharide.

The hydrogel forming polymer, optionally comprising gelatin, may bepresent in an amount of 300 to 700 mg/g (optionally 380 to 500 mg/g).The medium chain mono, di and/or tri-glycerides, may be present in anamount of 20 to 200 mg/g (optionally 40 to 80 mg/g). The solvent, forexample 2-(ethoxyethoxy)ethanol, may be present in an amount of 100 to250 mg/g (optionally 160 to 200 mg/g). The medium or long chain fattyacid mono- or di-ester or a combination thereof which does not compriseor is not a polyethyleneglycol ether or ester, for example glycerylmonooleate/dioleate, may be present in an amount of 80 to 200 mg/g(optionally 100 to 150 mg/g). The anionic surfactant, for example sodiumdodecyl sulphate, may be present in an amount of up to 100 mg/g or up to50 mg/g (optionally 10-70 mg/g, 15-60 mgm/g or 15-50 mg/g, preferably25-50 mg/g or 25-45 mg/g).

The composition or the core may comprise a hydrogel forming polymercomprising gelatin, optionally in an amount of 300 to 700 mg/g, the corefurther comprising medium chain mono, di and/or tri-glycerides,optionally in an amount of 20 to 200 mg/g, wherein the composition orcore further comprises the following components:

solvent, for example 2-(ethoxyethoxy)ethanol, optionally in an amount of100 to 250 mg/g;

a medium or long chain fatty acid mono- or di-ester or a combinationthereof which does not comprise or is not a polyethyleneglycol ether orester, for example glyceryl monooleate/dioleate, optionally in an amountof 80 to 200 mg/g; and

anionic surfactant, for example sodium dodecyl sulphate, optionally inan amount of up to 70 mg/g or up to 50 mg/g.

As will be recognised the composition or core further comprisescyclosporin.

The composition or the core may comprise:

a hydrogel forming polymer, for example which is, or comprises, gelatinin an amount of 300 to 700 mg/g;

cyclosporin in an amount of up to about 250 mg/g, for example 50 to 250mg/g;

medium chain triglycerides, for example Miglyol 810 in an amount of 20to 200 mg/g, optionally a solvent, for example 2-(ethoxyethoxy)ethanol,which when present is in an amount of 100 to 250 mg/g;

a surfactant comprising a medium or long chain fatty acid mono- ordi-ester or a combination thereof which does not comprise or is not apolyethyleneglycol ether or ester, for example glycerylmonooleate/dioleate, in an amount of 80 to 200 mg/g; and

anionic surfactant, for example sodium dodecyl sulphate, in an amount ofup to 60 mg/g or up to 50 mg/g, for example 10 to 50 mg/g, or optionally20 to 45 mg/g.

The composition or the core may comprise:

gelatin in an amount of 380-500 mg/g;

cyclosporin in an amount of 90-250 mg/g (optionally 90-200 mg/g or90-160 mg/g); and

caprylic/capric triglyceride in an amount of 40-80 mg/g;

2-(2-ethoxyethoxy) ethanol in an amount of 160-200 mg/g;

glyceryl monooleate and/or glyceryl dioleate in an amount of 100-150mg/g; and

SDS in an amount of 15-60 mg/g or 15-50 mg/g (optionally 25-50 mg/g or25-45 mg/g);

and

optionally D-sorbitol in an amount of 30-80 mg/g.

The composition or the core may comprise:

gelatin in an amount of 380-500 mg/g;

cyclosporin in an amount of 90-140 mg/g; and

caprylic/capric triglyceride in an amount of 40-80 mg/g;

2-(2-ethoxyethoxy) ethanol in an amount of 160-200 mg/g;

glyceryl monooleate and/or glyceryl dioleate in an amount of 100-150mg/g; and

SDS in an amount of 15-50 mg/g (optionally 25-50 mg/g or 25-45 mg/g);and

optionally D-sorbitol in an amount of 30-80 mg/g.

The composition or core may be a colloid. Where the composition or thecore is a colloid, the cyclosporin may be dissolved in the dispersephase of the colloid.

The composition or core may be a colloid; thus, the composition or coremay comprise a continuous phase and a disperse phase wherein thecontinuous phase comprises:

gelatin in an amount of 380-500 mg/g; and

optionally D-sorbitol in an amount of 30-80 mg/g;

the disperse phase comprises:

cyclosporin in an amount of 90-140 mg/g; and

caprylic/capric triglyceride in an amount of 40-80 mg/g;

and the composition further comprises:

2-(2-ethoxyethoxy)ethanol in an amount of 160-200 mg/g;

glyceryl monooleate and/or glyceryl dioleate in an amount of 100-150mg/g; and

SDS in an amount of 15-50 mg/g.

A colloidal composition or core comprising a continuous phasecomprising:

a hydrogel forming polymer matrix comprising gelatin in an amount of 300to 700 mg/g;

a disperse phase comprising:

cyclosporin in an amount of up to 200 mg/g; and

a medium chain tri-glyceride in an amount of 20 to 200 mg/g;

and the composition further comprising:

solvent in an amount of 100 to 250 mg/g;

surfactant (a first surfactant) being or comprising a medium or longchain fatty acid mono- or di-ester or a combination thereof which doesnot comprise is not a polyethyleneglycol ether or ester, for exampleglyceryl monooleate/dioleate; and

anionic surfactant (a second surfactant) in an amount of up to 50 mg/g.

In the embodiments above which refer to mg/g of a component, theconcentration is based upon the dry weight of the composition.

Suitably in the six compositions or cores described immediately above,the composition is a colloid comprising a disperse phase and acontinuous phase; wherein the disperse phase comprises cyclosporin,medium-chain triglyceride and medium or long chain fatty acid mono- ordi-ester surfactant; and the continuous phase comprises the hydrogelforming polymer (e.g. gelatin) and an anionic surfactant (e.g. SDS).

The invention includes within its scope compositions wherein the core isa colloid having a disperse phase and the continuous phase (matrixphase) of the colloid further includes dispersed particles of apharmaceutically active ingredient, for example microparticles ornanoparticles. The disperse phase and continuous phase may otherwise beas described elsewhere in this specification.

The composition of the invention and/or the core may be in the form of aminibead. It may be that the core is a minibead and the first coatingand, where applicable, the second coating in conjunction with the coreare in the form of a minibead. However, it may be possible for the coreto be a minibead and the composition not to be a minibead. Thecomposition may additionally comprise a multiplicity of minibeads. Hencethe invention contemplates a minibead with the features of thepharmaceutical compositions disclosed herein.

The composition or the minibead may have a largest cross sectionaldimension of a core of from about 0.01 mm to about 5 mm, for examplefrom 1 mm to 5 mm, as in the case of from 1 mm to 3 mm or 1 mm to 2 mm.The minibead may be spheroidal. The spheroidal minibeads may have anaspect ratio of no more than 1.5, for example from 1.1 to 1.5.

The composition of the invention may be for oral administration. Thecomposition may be formulated into a unit dosage form for oraladministration comprising from 0.1 mg to 1000 mg, optionally from 1 mgto 500 mg, for example 10 mg to 300 mg, or 25 to 250 mg suitably about25 mg, about 35 mg, about 37.5 mg, about 75 mg, about 150 mg, about 180mg, about 210 mg, about 250 mg or about 300 mg of cyclosporin. Suitablythe composition is in a multiple minibead unit dosage form selected frommultiple minibeads in, for example, soft or hard gel capsules, gelatincapsules, HPMC capsules, compressed tablets or sachets. The minibeadsmay be as described elsewhere herein.

The compositions described herein may be used to deliver cyclosporin Alocally to specific locations in the GIT, for example the solidcompositions described herein, may be adapted to provide release ofcyclosporin A in at least the colon. The compositions may be used toprovide the cyclosporin A locally in the GIT in a solubilised form,thereby providing high concentrations of cyclosporin in an active(available) form within the GIT where it acts to provide a therapeuticbenefit in a number of medical conditions, particularly conditionsaffecting the GIT as described in more detail herein, such as ulcerativecolitis. The release of cyclosporin A in an active form, for example asolubilised form, enables high concentrations of cyclosporin A to beabsorbed directly into the local tissues of the GIT, such as the colon.However, as described above, systemic exposure cyclosporin A has anumber of undesirable side effects. Therefore, a cyclosporin Acomposition which minimises systemic exposure to cyclosporin whilstmaintaining therapeutically beneficial concentrations in the tissues ofthe GIT would be desirable.

The major pathways of cyclosporin A metabolism in humans are viacytochrome P450 3A4 (CYP 3A4) and cytochrome P450 2J2 (CYP 2J2), withthree major metabolites being formed (two hydroxylated metabolites AM1,AM9 and one N-demethylated metabolite, AM4N). These metabolites haveminimal, if any immunosuppressive activity. Therefore minimising themetabolism of cyclosporin is desirable, because this minimises theformation of inactive metabolites and maximises the quantity ofcyclosporin available to interact locally with the tissues in the GIT.The primary metabolism of cyclosporin is via CYP 3A4, which is mainlyfound in the liver and the small intestine.

The total mass of CYP3A in the entire small intestine has been estimatedto be_1% of that in the liver. However, despite the relatively low massof CYP3A in the small intestine, enteric CYP3A can contributesignificantly, and in some cases equally, with hepatic CYP3A, to theoverall first-pass metabolism of several drugs including cyclosporin(Paine et al; Drug Metabolism and Disposition, Vol 34(5), 2006,880-885). CYP 3A4 expression in the colon is lower than in the smallintestine. Compositions which control the release of cyclosporin tolimit or inhibit release in the upper GI tract, for example, by usingone or more modified release coatings, may reduce enteric (i.e.non-systemic or “pre-systemic”) P450 metabolism. However, somemetabolism would still be expected resulting from the P450 expressed inthe tissues in the lower GIT.

It has been found that certain compositions described herein,particularly compositions which release cyclosporin in the lower GItract, especially in the colon, provide very low levels of cyclosporinmetabolism following oral administration of the composition. Thecompositions therefore maximise the amount of active (solubilised)cyclosporin available to interact with the tissues of the GI tractfollowing release of the cyclosporin from the composition. Withoutwishing to be bound by theory, it is thought that certain componentspresent in the composition, for example the medium chain or long chainfatty acid mono- or di-glyceride surfactant present in the composition,may act to inhibit cyclosporin metabolism by CYP 3A4 present in thetissues of the GI tract. When the composition is provided in a modifiedrelease format which prevents or inhibits release of cyclosporin in theupper GI tract, systemic absorption and metabolism of cyclosporin inliver is also minimised. Therefore, the modified release compositionsdescribed herein minimise both systemic and enteric metabolism ofcyclosporin. Low levels of cyclosporin metabolism may enable a lowerdose of cyclosporin to be administered whilst maintaining a therapeuticbenefit, thereby, widening the therapeutic window of the drug.

The relative degree of cyclosporin metabolism following oraladministration of a composition may be assessed by, for example,measuring the concentration of cyclosporin and the concentration ofcyclosporin metabolites present in a faecal sample collected from apatient following oral administration of a composition comprisingcyclosporin. As will be illustrated in the Examples, modified releasecompositions comprising cyclosporin and the surfactant (for exampleCapmul GMO-50) resulted in very low levels of cyclosporin metabolismcompared to a similar composition comprising a different surfactant(Cremophor). Compositions, particularly orally administeredcompositions, which exhibit low cyclosporin metabolism following releaseof cyclosporin from the composition, form a further aspect of theinvention.

Accordingly, there is provided a composition comprising cyclosporin A,wherein after oral administration of the composition to a human, themean concentration of cyclosporin A: the concentration of cyclosporin Ametabolites in a faecal sample from the human is greater than 12:1. Themean concentration of cyclosporin A:the concentration of cyclosporin Ametabolites in the faecal sample may be selected from: greater than19:1; greater than 24:1; greater than 31:1 and greater than 50:1. Themean concentration of cyclosporin A:the concentration of cyclosporin Ametabolites in the faecal sample may be selected from: from 20:1 to30:1; from 20:1 to 35:1; from 20:1 to 40:1; from 20:1 to 60:1; from 30:1to 50:1; and from 20:1 to 100:1. The mean concentration of cyclosporinA: the concentration of cyclosporin A metabolites in the faecal samplemay be selected from 12.5:1 to 90:1; from 13:1 to 85:1; from 15:1 to85:1; from 16:1 to 85:1; from 20:1 to 83:1; and from 65:1 to 79:1;optionally wherein the ratio is about 76:1.

The mean concentration of cyclosporin A: the concentration ofcyclosporin A metabolites in the faecal sample may be determined from afaecal sample collected from 12 to 28 hours after oral administration ofa single dose of the composition to the human. Alternatively the faecalsample may be collected after a more prolonged period of regular oraladministration of the composition to the human, after which thecyclosporin and metabolite concentrations in the faeces may have reacheda steady state, thereby reducing the variability in the measured ratioof cyclosporin:metabolites. For example, the faeces may be collected 4to 6 hours after the oral administration of the last dose of a dosageregimen wherein the cyclosporin composition is orally administered onceor twice per day for 2, 3 4, 5, 6 or 7 days. The faeces may, forexample, be collected after oral administration of 75 mg of cyclosporinonce or twice per day. Suitably, the faecal sample is collected 4 to 6hours after oral administration of the last dose of a dosing regimen ofthe composition; the dosing regimen comprising once or twice daily oraladministration of the composition to the human for seven days;optionally wherein the dosing regimen comprises once dailyadministration of the composition comprising 75 mg of cyclosporin A forseven days. In a further embodiment the composition comprisingcyclosporin is orally administered once per day for two days (forexample as a single 75 mg daily dose of cyclosporin) and the faecalsample is collected 4 to 6 hours after the last dose of the compositionon the second day.

The main metabolites of cyclosporin are the AM1, AM4N and AM9cyclosporin metabolites. The ratio of cyclosporin:cyclosporinmetabolites in the faecal sample is suitably the ratio ofcyclosporin:the total concentration of AM4N and AM9 cyclosporinmetabolites. The ratio of cyclosporin:cyclosporin metabolites in thefaecal sample may be the ratio of cyclosporin:the total concentration ofAM1, AM4N and AM9 cyclosporin metabolites.

The concentration of cyclosporin and its metabolites in the faeces maybe measured using any suitable analytical method, for examplechromatography and mass spectrometry as illustrated in the Examplessection.

Suitably the concentrations of cyclosporin and metabolites in the faecesare measured in faecal samples obtained from healthy male subjects so asto minimise the inter-patient variability in the measured values. Thefaecal samples are suitably obtained from healthy male subjects agedbetween 20 and 50 years, preferably weighing between 60 and 100 kg.Suitably the ratio is the arithmetic mean of the measured ratio ofcyclosporin:cyclosporin metabolites in a representative number ofsubjects, for example at least 4, 5, 6, 7, 8, 9 or 10 subjects.Generally at least 4 subjects would be a sufficient number to provide arepresentative mean ratio.

The composition comprising cyclosporin A may be any compositioncomprising cyclosporin A which provides a ratio ofcyclosporin:cyclosporin metabolites which is greater than 12:1 (orwithin any of the ranges described above in relation to this aspect ofthe invention). The composition is suitably a composition comprisingcyclosporin A, wherein the composition releases cyclosporin A in asolubilised form when the composition is placed in an aqueousdissolution medium. By “solubilised” in meant that the cyclosporin inreleased in an active form, for example in a dissolved form such as asolution, when the composition is placed in an aqueous dissolutionmedium, for example an aqueous environment encountered in the lower GITtract, particularly in the colon, following oral administration of thecomposition.

The composition may be, or comprise, cyclosporin A that is partially orcompletely dissolved in dissolved in a lipophilic substance. Thelipophilic substance may be or comprise an oil, or a surfactant in whichthe cyclosporin is at least partially, or preferably fully dissolved.Suitable oils and surfactants which may be used include, but are notlimited to any of the the oils and surfactants described herein.

In one embodiment the cyclosporin may be dissolved or dispersed in a lowmelting point lipophilic substance, suitably a substance with a meltingpoint in the range of 30 to 70° C. Suitably the hydrophobic material isa wax like solid with a melting point in the range of 30 to 60° C.,particularly suitable are lipophilic waxy material which are solid atroom temperature, but which melt or soften at temperatures in the rangeof 30 to 50° C., or more preferably 30 to 40° C. The lipophilic materialmay be selected from one or more of unsaturated alcohols, hydrogenatedalcohols, fatty acids, fatty acid esters, fatty acid amides, fatty acidmono- di- or triglycerides, polyethoxylated fatty acids andpolyethyoxylated fatty acid esters, cholesterol derivatives and waxes.The wax may be a suitable animal or plant derived wax, for examplecarnauba wax or a synthetic wax such as paraffin wax. The lipophilicmaterial may comprise a wax, a saturated of unsaturated fatty acid (forexample palmitic, stearic, myristic, lauric, laurylic, or oleic acid),or a derivative thereof for example a mono-, di-, or triglyceride or apolyethylene glycol ester thereof. The lipophilic material comprisingthe dissolved or dispersed cyclosporin is suitably used in the form of aparticulate composition, for example as a granule composition. Suitablythe lipophilic substance comprising the dissolved or dispersedcyclosporin is itself dispersed, in a suitable carrier matrix. Forexample, granules comprising the the lipophilic substance and dissolvedor dispersed cyclosporin are dispersed in a suitable carrier matrix. Thecarrier matrix may be a modified release matrix material, particularly amodified release polymer matrix. A modified release matrix may providedelayed or sustained release of the cyclosporin from the matrix, therebyproviding a modified release composition. The carrier matrix is suitablya hydrophilic material. The matrix material may be, or comprise, acrylicor methacrylic acid polymers or copolymers, alkylvinyl polymers,hydroxyalkyl celluloses, carboxyalkyl celluloses, polysaccharides,dextrins, pectins, starches, starch derivatives, or natural or syntheticgums for example an alginate. The carrier matrix may be a hydrogelforming polymer such as those described herein, including gelatin.

The composition may comprise cyclosporin A and a surfactant. Suitablythe surfactant is or comprises a medium chain or long chain fatty acidmono- or di-glyceride or a combination thereof as described herein.Suitably, the in this embodiment the weight ratio of cyclosporin tomedium chain or long chain fatty acid mono- or di-glyceride or acombination is from about 3:1 to about 1:3, for example about 3:1 toabout 1:2, or about 2.5:1 to about 1:1.8, about 1.5:1 to 1:1.5, about1.2:1 to 1:1.2, about 1.2:1, about 1:1, or about 1:1.2. Suitably thecyclosporin composition in this embodiment further comprises an oilphase. The oil phase may be any suitable hydrophobic oil, for example anoil having a low HLB value (for example HLB less than 10). Particularlythe oil phase may comprise any of the oil phases described herein, forexample the oil phase may be or comprise triglycerides, for example amedium chain triglyceride. The weight ratio of oil to surfactant may befor example 12:1 to 1:5, for example from about 5:1 to about 1:5, fromabout 3:1 to about 1:2, from about 3:1 to about 1:1 or from about 2.5:1to 1.5:1. The composition may further comprise a solvent. The solvent issuitably an organic solvent in which cyclosporin is soluble. Moreparticularly suitable solvents include those in which both cyclosporinand the oil phase (when present) are soluble. For example the solventmay comprise 2-(ethoxyethoxy)ethanol. The cyclosporin may be partiallyor fully dissolved in the composition. Accordingly, the cyclosporin maybe substantially dissolved in the composition. Suitably the cyclosporinis fully dissolved in the composition.

The composition is suitably formulated such that release of cyclosporinin the upper GI Tract, for example in the duodenum and jejunum, isminimised so as to minimise the systemic absorption of cyclosporin andboth hepatic and enteric P450 metabolism. Accordingly a particularcomposition is a modified release composition. Suitably the release ofcyclosporin from the composition is minimised for the first 4 hoursafter oral administration such that the composition can pass through theduodenum and jejunum and into the ileum before releasing large amountsof cyclosporin. Preferably the composition releases the majority (forexample at least 50%) of the cyclosporin into the colon. Suitably thecomposition releases less than 40% (for example less than 35%, or lessthan 30%) of the cyclosporin from the composition 4 hours when measuredin the two stage dissolution test described herein. Suitably thecomposition releases less than 15% (for example 0 to 10%) of thecyclosporin A after 2 hours; and releases 10% to 40% (for example 10% to35%, or suitably 15% to 35%) of the cyclosporin A at 4 hours, whenmeasured in the two stage dissolution test.

The composition may be formulated to provide the desired modifiedrelease profile by, for example, use of any of the coatings describedherein, in particular coatings which are adapted to release cyclosporinin at least the colon. Suitable coatings include, for example, amodified release coating comprising a pH independent polymer such asethyl cellulose. The coating may also comprise a first coatingcomprising a water soluble cellulose ether such as HPMC, as describedherein. Other modified release coatings are also contemplated includingbut not limited to enteric coating systems and other delayed releasecoatings. Generally the modified release coatings comprise a polymericcoating.

In this aspect of the invention, when the composition comprisescyclosporin A and a surfactant which is or comprises a medium chain orlong chain fatty acid mono- or di-glyceride or a combination thereof asdescribed herein, the cyclosporin and surfactant are suitably dispersedwithin in matrix. The cyclosporin is released from the matrix when thecomposition is placed in an aqueous environment, for example as would befound in the lower GI tract such as in the colon. Suitable matrixmaterials which may be used to disperse the cyclosporin and surfactantmay be any of the matrix materials described herein, for example thosedescribed under “Composition” in the detailed description. In certainembodiments the matrix material may be selected such that the matrixitself modifies the release of cyclosporin from the composition asdescribed in more detail in the detailed description of the invention.In such compositions it may be possible to achieve the desiredinhibition of release of cyclosporin in the first 4 hours following oraladministration without the need for additional modified releasecoating(s). In other embodiments the matrix material may be coated withone or more of the modified release coatings, such as those describedherein to provide the required cyclosporin release profile. In aparticular, in this aspect of the invention the matrix is a hydrogelpolymer, as described herein, more particularly the matrix is orcomprises gelatin.

In a preferred embodiment in this aspect of the invention, thecomposition comprising cyclosporin may be any of the cyclosporincompositions described herein which comprise a surfactant wherein thesurfactant is, or comprises, a medium chain or long chain fatty acidmono- or di-glyceride or a combination thereof.

The cyclosporin compositions described herein, more particularly themodified release compositions described herein, provide pharmacokinetic(PK) properties which minimise systemic exposure to cyclosporin comparedto, for example, intravenous administration of cyclosporin and/or oraladministration of instant release cyclosporin compositions such asNeoral™. The following paragraphs describing suitable PK properties ofthe compositions are applicable to any of the cyclosporin compositionsdescribed herein.

The composition may provide a low systemic whole blood exposure tocyclosporin following oral administration of the composition. Thecomposition may provide a mean whole blood cyclosporin A AUC_(0-inf) ofless than about 450 ng·hr/ml, less than about 350 ng·hr/ml, or less thanabout 300 ng·hr/ml after oral administration of the composition as asingle dose containing 75 mg cyclosporin A to a human in a fasted state,or an AUC_(0-inf) directly proportional thereto for a total dose otherthan 75 mg. For example, the composition may provide a mean whole bloodcyclosporin A AUC_(0-inf) of from about 140 to about 420 ng·hr/ml, forexample from about 150 to about 300 ng·hr/ml after oral administrationof the composition as a single dose containing 75 mg cyclosporin A to ahuman in a fasted state, or an AUC_(0-inf) directly proportional theretofor a total dose other than 75 mg.

A high peak blood concentration of cyclosporin A (C_(max)) may result inundesirable side effects and potentially reduce the therapeutic windowavailable for a composition containing cyclosporin A. Accordingly, thecompositions suitably provide a low C_(max). The composition comprisingcyclosporin A, may provide a mean maximum whole blood concentration ofcyclosporin A (C_(max)) of less than 100 ng/ml. The composition may forexample provide a C_(max) of from about 15 to about 60 ng/ml, forexample about 20 to 50 ng/ml, wherein in each case the C_(max) is thatmeasured after oral administration of the composition as a single dosecontaining 75 mg of cyclosporin A to a human in a fasted state, or aC_(max) directly proportional thereto for a total dose other than 75 mg.

The time taken to reach maximum whole blood concentration (Tmax) of thecyclosporin A suitably occurs between about 3 and about 10 hours afteroral administration of the composition as a single dose to a human in afasted state. The Tmax may occur between about 4 hours and about 10hours, or between about 4 hours and about 8 hours, or between about 5and about 6 hours after oral administration of the composition. Forexample a T_(max) at about 5 hours, about 5.5 hours or about 6 hours.

An IV dose of 2 to 4 mg/kg/day cyclosporin is known to be efficacious inthe treatment of ulcerative colitis patients (Lichtiger et al N. Engl JMed 1994; 330: 1841-1845). An IV dose of 2 mg/kg is approximates to acyclosporin dose of approximately 150 mg (assuming an average weight ofabout 75 kg). As illustrated in the examples section it has been foundthat the AUC resulting from IV administration of cyclosporin asSandimmun™ is significantly higher than the AUC resulting from themodified release compositions comprising the surfactant. IVadministration of cyclosporin results in effectively 100% systemicbioavailability. Accordingly, a comparison of the AUC for IVadministration with the orally administered cyclosporin compositionsdescribed herein enable the absolute oral bioavailability (F %) to bedetermined. As illustrated in the examples, the cyclosporin compositionsdescribed herein provide a low absolute oral bioavailability. The F % iscalculated by calculating the relative % of the AUC following oraladministration relative to the AUC observed following IV administrationof 2 mg/kg Sandimmun™. As will be realised compensation for the actualdose of cyclosporin needs to be accounted for when calculating the F %.For example if the AUC for oral administration is that measuredfollowing a single dose of 75 mg of cyclosporin, the relative % needs tobe multiplied by 2 to compensate for the fact that the effective totalIV dose was 150 mg cyclosporin. Similarly, if a 37.5 mg dose ofcyclosporin is administered orally, the relative % needs to bemultiplied by 4.

The composition comprising cyclosporin A may provide a cyclosporin Aabsolute bioavailability following oral administration of thecomposition is less than 15%, for example less than 10%; optionallywherein the absolute bioavailability is from 0.5% to 15% suitably from1% to 10%.

Suitably the composition releases the cyclosporin in at least the colon.The composition may also release cyclosporin in other parts of the GItract for example in the duodenum, jejunum and/or ileum. Suitablyhowever, release of cyclosporin in the upper GI tract such as theduodenum and jejunum is minimised so as to reduce systemic exposure tocyclosporin A and/or reduce P450 metabolism of the drug. The releaseprofile of cyclosporin A from the composition may be assessed bymeasuring the release in an in vitro dissolution test. The compositioncomprising cyclosporin may release less than 15% (for example 0 to 10%)of the cyclosporin A after 2 hours; releases 10% to 40% (for example 10%to 35%, or suitably 15% to 35%) of the cyclosporin A at 4 hours; andreleases from about 30% to 70% (for example 40% to 70%) of thecyclosporin A between 4 hours and 12 hours, when measured in a two stagedissolution test using a USP Apparatus II with a paddle speed of 75 rpmand a dissolution medium temperature of 37° C.; wherein for the first 2hours of the dissolution test the dissolution medium is 750 ml of 0.1 NHCl, and at 2 hours 250 ml of 0.2M tribasic sodium phosphate containing2% SDS is added to the dissolution medium and the pH is adjusted to pH6.8 (herein referred to as “the two-stage dissolution test).

The composition may release less than 20% of the cyclosporin A after 2hours; releases 10 to 40% of the cyclosporin A at 4 hours; and releasesat least 60% of the cyclosporin A at 12 hours, when measured in the twostage dissolution test. The composition may release less than 10% of thecyclosporin A after 2 hours; releases 10 to 30% of the cyclosporin A at4 hours; and releases at least 50% of the cyclosporin A at 12 hours,when measured in the two stage dissolution test. The composition mayrelease from about 30 to about 75% of the cyclosporin A between 4 hoursand 12 hours in the two stage dissolution test, for example thecomposition releases from about 40 to about 75%, particularly from about45 to 70% of the cyclosporin A between 4 hours and 12 hours in the twostage dissolution test. The composition may release less than 15% (forexample 0 to 10%) of the cyclosporin A after 2 hours; releases 10% to40% (for example 10% to 35%, or suitably 15% to 35%) of the cyclosporinA at 4 hours; and releases from about 25% to 70% (for example 40% to70%) of the cyclosporin A between 4 hours and 12 hours in the two stagedissolution test.

It is to be understood that any of the individual PK parameters and/orin-vitro or other release profiles described herein may be combined withthe compositional features of the cyclosporin compositions describedherein, for example relating to any one or a combination of AUC; Cmax;Tmax; cyclosporin A concentration in the luminal contents; cyclosporin Aconcentration in the GI tract tissue; the ratio of cyclosporin A in theluminal contents:cyclosporin A in GI tract tissues; the ratio of theconcentration of cyclosporin A:the concentration of cyclosporin Ametabolites in collected faeces; the concentration of cyclosporin A inintracolonic faeces:the concentration of cyclosporin A in colonictissue; or the concentration of cyclosporin A in colonic tissue. By wayof a non-limiting example of such a combination of features compositioncomprising cyclosporin A, provides a mean concentration of cyclosporinA: the concentration of cyclosporin A metabolites in a faecal samplefrom the human of greater than 12:1 after oral administration of thecomposition to the human; and wherein the composition provides anAUC_(0-inf) of less than about 450 ng·hr/ml, (e.g. from about 140 toabout 420 ng·hr/ml) after oral administration of the composition as asingle dose containing 75 mg cyclosporin A to a human in a fasted state,or an AUC_(0-inf) directly proportional thereto for a total dose otherthan 75 mg. Optionally this composition may release 0 to 10% of thecyclosporin A after 2 hours; 10% to 35% of the cyclosporin A at 4 hours;and releases from about 40% to 70% of the cyclosporin A between 4 hoursand 12 hours, when measured in the two stage dissolution test.

In another embodiment the composition releases 0 to 10% of thecyclosporin A after 2 hours; and releases from 50 to 100% of thecyclosporin A after 12 hours, when measured in the two stage dissolutiontest. In another embodiment the composition releases less than 20% ofthe cyclosporin A after 2 hours; releases 5 to 40% of the cyclosporin Aat 4 hours and releases at least 50% of the cyclosporin A at 12 hours,when measured in the two stage dissolution test.

The cyclosporin compositions described herein are expected to providesimilar or higher levels of cyclosporin A in the colonic tissue comparedto IV administration of Sandimmun™, but with a higher intracolonicfaecal concentration of cyclosporin A as a result of the local releaseof the cyclosporin directly into the colon. The relatively high localconcentration of cyclosporin in the colon is expected to providebeneficial therapeutic effects.

The composition comprising cyclosporin A described herein, may provide aratio of the mean concentration of cyclosporin A present in intracolonicfaeces:the mean concentration of cyclosporin A present in colonic tissuein an adult human patient after oral administration of the compositionof from about 50:1 to about 500:1, optionally from about 80:1 to about300:1, or optionally about 100:1 to about 250:1; wherein theconcentration of the cyclosporin A is measured in samples of theintracolonic faeces and the colonic tissue taken substantiallysimultaneously 4 to 6 hours after oral administration of the last doseof a once daily oral dosing regimen of the composition, the dosingregimen comprising once daily oral administration of the composition forseven days. Optionally the once daily oral dosing regimen of thecomposition provides a single daily dose of 75 mg cyclosporin A.However, other doses may be administered for example any of thecyclosporin doses described herein including but not limited to 37.5 mgor 150 mg once per day. Optionally the dosage regimen may be a twicedaily dosage regimen for seven days, for example 37.5 mg twice per day,75 mg twice per day or 150 mg twice per day.

Reference herein to a sample being taken “substantially simultaneously”means that the samples are obtained close to the same time point, forexample the colonic tissue and/or intracolonic faeces and/or bloodsamples are taken within about 2 hours, 1 hour or 30 minutes of eachother, suitably the samples are all taken at the same time point.

In contrast to the compositions according to the invention, IVadministration cyclosporin as Sandimmun™ results in a lower ratio of themean concentration of cyclosporin A present in intracolonic faeces:themean concentration of cyclosporin A present in colonic tissue. Asillustrated in the Examples, when patients were treated with Sandimmun®IV (2 mg/kg) administered as an infusion over 24 hours (2 mg/kg/day) aratio of about 3:1 was observed.

Accordingly, the orally administered cyclosporin compositions describedherein are expected to provide a relatively high colonic tissueconcentrations compared to IV administration of an equivalent dose ofcyclosporin (e.g. as Sandimmun™ IV).

The composition comprising cyclosporin A may provide a concentration ofcyclosporin A in colonic tissue of at least 250 ng/g following oraladministration of the composition to a human, for example at least 300,350, 400 or 450 ng/g. Accordingly, the composition may provide acyclosporin A concentration of from about 250 to 6000 ng/g, for examplefrom 400 ng/g to 6000 ng/g, from 500 to 5000 ng/g, from 600 to 4000ng/g, or from 600 to 2000 ng/g. Particularly the composition provides acyclosporin A concentration in colonic tissue of from about 1000 toabout 1500 ng/g, for example about 1200 ng/g. Suitably the compositionis orally administered to provide a daily dose of cyclosporin A withinthe ranges described herein, suitably a total daily dose in the range of15 to 300 mg cyclosporin. Optionally the composition provides a dose maybe 37.5 mg, 75 mg or 150 mg cyclosporin A once or twice per day.

The colonic tissue samples described above may be obtained usingconventional methods, for example by taking a tissue sample duringendoscopy as described in the examples herein.

As described above, compositions comprising cyclosporin A and asurfactant, wherein the surfactant comprises, or is a medium chain orlong chain fatty acid mono- or di-glyceride or a combination thereof mayinhibit P450 metabolism of cyclosporin A. Such compositions may beuseful for the preparation of modified release of cyclosporin in thelower GI tract as described above. Also contemplated are suchcompositions for administration of cyclosporin to any part of the GItract, for example duodenum or jejunum as, for example, an instantrelease composition. The compositions may reduce the rate and or extentof cyclosporin metabolism and thereby maximise the amount of cyclosporinin the GI tract.

According to a further feature of the invention there is provided acomposition comprising cyclosporin A and a surfactant, wherein thesurfactant comprises, or is a medium chain or long chain fatty acidmono- or di-glyceride or a combination thereof. Suitably the compositiondoes not comprise or is not a polyethyleneglycol ether or ester.Suitably the surfactant is present in an amount of at least 6% by weightof the composition, for example at least 10%, at least 15% or at least20% by weight of the composition. Optionally the surfactant is presentin an amount of from 10 to about 50% by weight.

The composition according to this aspect of the invention may furthercomprise an oil phase, for example any of the oil phases describedherein.

The cyclosporin may be partially or completely dissolved in thecomposition. Suitably the cyclosporin is completely dissolved in thecomposition.

A particular composition comprises:

(i) 10 to 60 parts cyclosporin A;

(ii) 5 to 40 parts of a medium chain fatty acid triglyceride, forexample a caprylic/capric triglyceride;

(iii) 10 to 50 parts of the surfactant; and

(iv) 0 to 60 parts solvent;

wherein all parts are parts by weight and the sum of the parts(i)+(ii)+(iii)+(iv)=100.

Another composition comprises:

(i) 10 to 40 parts cyclosporin A;

(ii) 5 to 25 parts of a medium chain fatty acid triglyceride, acaprylic/capric triglyceride;

(iii) 15 to 30 parts of the surfactant; and

(iv) 10 to 60 parts solvent (optionally 20 to 40 parts or 25-30 partssolvent), for example 2-(2-ethoxyethoxy)ethanol);

wherein all parts are parts by weight and the sum of the parts(i)+(ii)+(iii)+(iv)=100.

Optionally in this aspect the surfactant is selected from glycerylcaprylate, glyceryl caprate, glyceryl monooleate, glyceryl dioleate andglycerol monolinoleate, or a combination thereof.

The cyclosporin compositions according to this aspect may beadministered orally, for example to provide an instant releasecomposition. Also contemplated is the administration of the compositionto the GI tract rectally, for example in the form of an enema orsuppository. Other routes of administration of the composition are alsocontemplated, for example the composition may be administered directlyto the GIT, by for example intra-duodenal administration, intra-jejunalor intra-ileal administration. Such routes of administration enable thecomposition to bypass the stomach (and optionally other parts of the GItract) for delivery to specific points in the lower GI tract. Theseroutes of administration may be achieved using for example suitabletubing with an exit at the desired location within the GI tract.Suitably the tubing is inserted orally or nasally into the GI Tract.Alternatively, administration may be achieved by gastric tubing, orcontinuous or discontinuous percutaneous endoscopic gastrostomy (PEG)tubing. PEG is an endoscopic medical procedure in which a tube (PEGtube) is passed into a patient's stomach through the abdominal wall.This method of administration may be particularly suitable for patientsthat cannot take the drug orally due to for example dysphagia orsedation.

A further aspect of the invention provides a composition describedherein for use as a medicament. The composition may comprise at leastone further active ingredient, for example at least one furtherimmunosuppressant. In particular there is provided a composition for usein the treatment, e.g. prevention, of a condition of the GIT. Thecomposition may be for use in the treatment of an inflammatory boweldisease, irritable bowel syndrome, Crohn's disease, ulcerative colitis,celiac disease, graft-versus-host disease, gastrointestinalgraft-versus-host disease, gastroenteritis, duodenitis, jejunitis,ileitis, peptic ulcer, Curling's ulcer, appendicitis, colitis,pseudomembraneous colitis, diverticulosis, diverticulitis, pouchitis,collagenous colitis, macorscopic colitis, diarrheal colitis,endometriosis, colorectal carcinoma and adenocarcinoma. The compositionmay also be for use in the treatment of proctitis. The composition maybe for use in the prevention or treatment of primary sclerosingcholangitis, familial adenomatous polyposis, or perinanal Crohn's,including perianal fistulae.

In embodiments where the pharmaceutical composition does not comprise asecond coating, the composition may be for use in the treatment ofconditions that affect the small intestine. Such compositions may beable to treat conditions selected from celiac disease, GVHD or Crohn'sdisease.

The invention additionally provides a method for administeringcyclosporin to a subject, comprising orally administering to the subjecta composition described herein. The method may be performed in thetreatment, e.g. prevention, of disease. The subject may be a mammal, inparticular a human. Also provided is a method for treating a conditionof the GI tract in a subject, preferably a human, in need thereofcomprising orally administering to the mammal a therapeuticallyeffective amount of a composition described herein. Conditions of the GItract which may be treated or prevented include the conditions disclosedherein.

A further aspect of the invention provides the use of a compositiondescribed herein for use in the manufacture of a medicament for thetreatment, e.g. prevention, of a condition of the GIT. Conditions of theGI tract include those disclosed herein.

The invention also contemplates a method of treating a conditionselected from inflammatory bowel disease, irritable bowel disease,Crohn's disease, ulcerative colitis, celiac disease, graft vs hostdisease, gastrointestinal graft-versus-host disease, gastroenteritis,duodenitis, jejunitis, ileitis, peptic ulcer, Curling's ulcer,appendicitis, colitis, pseudomembraneous colitis, diverticulosis,diverticulitis, collagenous colitis, endometriosis, colorectal carcinomaand adenocarcinoma, wherein the method comprises administering apharmaceutical composition of the invention.

In another aspect the invention provides a method of treating conditionsthat affect the small intestine, wherein the method comprisesadministering a composition of the invention which does not comprise asecond coating. The conditions of the small intestine may be selectedfrom celiac disease, GVHD or Crohn's disease.

In an aspect of the invention there is provided a process for making aliquid composition, the process comprising mixing an oil phase with anaqueous phase comprising a hydrogel forming polymer, wherein the oilphase has cyclosporin in solution and comprises a surfactant which ismedium chain or long chain fatty acid mono- or di-glyceride or acombination thereof, wherein the surfactant does not comprise or is nota polyethyleneglycol ether or ester.

Optionally, the oil phase and the aqueous phase are mixed in an oilphase to aqueous phase ratio of from 1:2 to 1:12, optionally 1:4 to1:10, 1:4 to 1:8, for example 1:5 or 1:7.

The process may further comprise the step of causing the emulsion tosolidify.

The process may further comprise the step of:

coating a core with a coating comprising HPMC wherein the weight gaindue to the coating is from 0.5% to 20% of the weight of thepharmaceutical composition. The core may comprise a pharmaceuticallyactive ingredient and may be a core as described in this specification.

An additional advantage of the present application may be that acomposition dissolved in a dissolution medium yields a uniform dropletsize with low polydispersivity compared to formulations with a differentfirst surfactant.

Accordingly, there is provided a composition comprising cyclosporin, ahydrogel forming polymer matrix, a surfactant and an oil phase beingdispersed in the hydrogel forming polymer matrix, wherein the surfactantis or comprises a medium chain or long chain fatty acid mono- ordi-glyceride or a combination thereof and does not comprise or is not apolyethyleneglycol ether or ester,

wherein the composition releases droplets with a uniform size.Optionally the droplets may have low polydispersity. Optionally theuniform size may be a droplet size of from about 1 nm to about 350 nm.

The droplet size may be selected from: from about 20 nm to about 350 nm;from about 20 nm to about 300 nm; from about 20 nm to about 250 nm fromabout 100 nm to about 350 nm; from about 100 nm to about 300 nm; fromabout 100 nm to about 250 nm; from about 100 nm to about 200 nm; fromabout 150 nm to about 250 nm; from about 150 nm to about 200 nm; fromabout 150 nm to about 350 nm; and from about 150 nm to about 300 nm.Preferably, the droplet size may be selected from: from about 20 nm toabout 250 nm; from about 100 nm to about 250 nm; and from about 100 nmto about 200 nm.

The size of the droplets may be measured using dynamic light scattering.The dynamic light scattering experiments were carried out by analysis ofa liquid medium arrived at as follows. Minibeads of the invention (0.5g) comprising Capmul GMO-50 as the first surfactant were added to abeaker containing 50 g of deionised water. The beaker contents weremixed at 250 rpm and at 37° C. throughout the study. Samples of thebeaker contents were taken at 0, 1, 2, 3, 4, 5, 6 and 24 hours. Samplesof the beaker contents were filtered 0.65 μm pore size filters (MerckMillipore Ultrafree-CL Centrifugal Filter). The particle size and zetapotential was measured using a Malvern Nano-Zetasiser.

For certain active ingredients it may be desirable to limit or delayrelease of the active from the composition until the composition haspassed through the stomach and upper GI tract. The compositions of theinvention comprising a second coat may be particularly suitable for suchapplications. The second coat acts to delay release from thecomposition, whilst the presence of the coating of the invention (e.g.HPMC) increases the amount of active released when the compositionreleases the active in the lower GI tract. The period of delay to therelease of the active as a result of the presence of the second coatingcan be tailored by appropriate selection of the nature or amount ofsecond coating used. For a given second coating material a higher weightgain of coating will generally increase the time period betweenadministration of the composition and release of the active. Thecompositions of the invention can therefore be used to provide highlevels of release of active agent at very specific parts of the GI tractto provide, for example, topical treatment to diseased tissue within theGI tract. Such delayed release compositions may be particularlybeneficial when the active has undesirable side effects which may arisefrom systemic absorption higher in the GI tract.

Included in this description by reference are the subject matters of theappended claims. The description is therefore to be read together withthe claims and features mentioned in the claims are applicable to thesubject matters of the description. For example, a feature described ina process claim is applicable also to products mentioned in thedescription, where the feature is manifested in the product. Forexample, a feature mentioned in a product claim is applicable also torelevant process subject matters contained in this description.Similarly, a feature mentioned in the description in the context of aprocess is applicable also to products mentioned in the description,where the feature is manifested in the product. Also, a featurementioned in the description in the context of a product is applicablealso to relevant process subject matters contained in this description.

BRIEF DESCRIPTION OF THE DRAWINGS

Embodiments of the invention are further described hereinafter withreference to the accompanying drawings, in which:

FIGS. 1A-1I are images showing crystal formation over time of acomparative composition.

FIGS. 2A-2I are images showing crystal formation over time of acomposition of the invention comprising Capmul GMO-50 (glycerylmonooleate/dioleate) as the surfactant (the first surfactant).

FIGS. 3A-3J are images showing crystal formation over time of acomposition of the invention comprising Capmul MCM (glycerylcaprylate/caprate) as the surfactant (the first surfactant).

FIGS. 4A-4J are images showing crystal formation over time of acomposition of the invention comprising Maisine 35-1 (glycerolmonolinoleate) as the surfactant (the first surfactant).

FIG. 5 is a graph plotting % of cyclosporin released against time over24 hours and showing the release profiles of minibeads of Example 2.

FIG. 6 is a graph plotting % of cyclosporin released against time over24 hours and showing the release profiles of minibeads of ComparativeExample 1.

FIG. 7 is a graph plotting % of cyclosporin released against time over24 hours in deionised water and showing the release profiles ofminibeads of Example 6a.

FIG. 8 is a graph plotting % of cyclosporin released against time over24 hours in the two-stage dissolution test showing the release profilesof beads of Example 4a, specifically those of Table 11.

FIG. 9 is a graph showing the droplet size and zeta potential of acoated composition of the invention when the composition has beendissolved in deionised water.

FIG. 10 shows the dosing schedule used in the clinical trial describedin Example 9.

FIG. 11 shows the median whole blood cyclosporin concentration timeprofiles (linear linear) on day 1 in those subjects treated with CyCol®in the clinical trial described in Example 9.

FIG. 12 the median whole blood cyclosporin concentration time profiles(linear linear) on day 7 in those subjects treated with CyCol® in theclinical trial described in Example 9. In FIGS. 11 and 12, BID=twicedaily; OD=once daily. Values below the lower limit of quantification(<0.2 ng/mL) are presented as equal to zero.

FIGS. 13A-13B show the mean whole blood concentration of cyclosporin Aobtained from the comparative clinical trial of Comparative Example 10on linear and log scales.

FIG. 14 shows the ratio of cyclosporin A to the concentration of the(AM4N+AM9) cyclosporin metabolites measured in the faecal samples foreach of the tested formulations in Example 9 and Comparative Example 10.

FIG. 15 shows the in-vitro release profiles of the compositions used inExample 9 and Comparative Example 10. In FIG. 14 “PK fast”, “PK medium”and “PK slow” refer to the Fast Release Formulation, Formulation I andFormulation II used in Comparative Example 10. “CyCol 2014” refers tothe formulation used in Example 9.

FIGS. 16A-221 show photomicrographs of the emulsions described inExample 11 comprising surfactants in an aqueous phase at specified timepoints.

DETAILED DESCRIPTION

A mono-glyceride or di-glyceride of the present invention may compriseone glycerol esterified to one fatty acid or one glycerol esterified totwo fatty acids the fatty acids may be the same or different, ordinarilythe fatty acids will be the same. The surfactant of the invention is asurfactant that does not comprise or is not a polyethyleneglycol etheror ester; by this it is meant that there is no polyethyleneglycolcomponent bonded to the surfactant molecule by an ether or esterlinkage. For example a pegylated fatty acid glyceride such as oleoylmacrogol-6 glycerides (commercially available as Labrafil M1944CS). Itis possible that a commercial surfactant of the invention is suppliedwith a small amount of polyethyleneglycol (PEG) contained within thesupplied surfactant composition. The use of such commercial formulationsof surfactants which contain non-bonded PEG, put another way free PEG)are not excluded by the limitation that the surfactant does not compriseor is not a polyethyleneglycol ether or ester.

Reference to “cyclosporin” herein is a reference to cyclosporin-A (alsoknown as cyclosporine and the INN ciclosporin. It is contemplated thatother forms of cyclosporin may be used in the compositions describedherein, for example cyclosporin-B, -C, -D or -G and derivatives or prodrugs of any thereof.

The term “treatment”, and the therapies encompassed by this invention,include the following and combinations thereof: (1) reducing the risk ofor inhibiting, e.g. delaying, initiation and/or progression of, a state,disorder or condition; (2) preventing, e.g. reducing the risk of, ordelaying the appearance of clinical symptoms of a state, disorder orcondition developing in a patient (e.g. human or animal) that may beafflicted with or predisposed to the state, disorder or condition butdoes not yet experience or display clinical or subclinical symptoms ofthe state, disorder or condition; (3) inhibiting the state, disorder orcondition (e.g., arresting, reducing or delaying the development of thedisease, or a relapse thereof in case of maintenance treatment, of atleast one clinical or subclinical symptom thereof); and/or (4) relievingthe condition (e.g. causing regression of the state, disorder orcondition or at least one of its clinical or subclinical symptoms).Where the composition of the invention is used in the treatment of apatient, treatment contemplates any one or more of: maintaining thehealth of the patient; restoring or improving the health of the patient;and delaying the progression of the disorder. The benefit to a patientto be treated may be either statistically significant or at leastperceptible to the patient or to the physician. It will be understoodthat a medicament will not necessarily produce a clinical effect inevery patient to whom it is administered, and this paragraph is to beunderstood accordingly. The compositions and methods described hereinare of use for therapy and/or prophylaxis of disease.

The treatments may include maintenance therapy of patients who havesuffered a disorder and whose condition has subsequently improved, e.g.because of treatment. Such patients may or may not suffer a symptomaticdisorder. Maintenance therapy aims to arrest, reduce or delay (re-)occurrence or progression of a disorder.

“Effective amount” means an amount sufficient to achieve the desiredtreatment, e.g. result in the desired therapeutic or prophylacticresponse. The therapeutic or prophylactic response can be any responsethat a user (e.g., a clinician) will recognise as an effective responseto the therapy. It is further within the skill of one of ordinary skillin the art to determine appropriate treatment duration, appropriatedoses, and any potential combination treatments, based upon anevaluation of therapeutic or prophylactic response.

The terms “dry” and “dried” as applied to compositions or compositionsof the disclosure may each include reference to compositions orcompositions containing less than 5% free water by weight, e.g. lessthan 1% free water by weight. Primarily, however, “dry” and “dried” asapplied to compositions of the disclosure mean that the hydrogel presentin the initial solidified composition has dried sufficiently to form arigid composition. Where a solid colloid is referred to this also refersto a dried colloid according to the definition herein.

Ingredients and excipients of the described compositions are suitablefor the intended purpose. For example, pharmaceutical compositionscomprise pharmaceutically acceptable ingredients.

If not otherwise stated, ingredients, components, excipients etc. of thecompositions of the invention are suitable for one or more of theintended purposes discussed elsewhere herein.

For the avoidance of doubt, it is hereby stated that the informationdisclosed earlier in this specification under the heading “Background”is relevant to the invention and is to be read as part of the disclosureof the invention.

Where the invention is referred to as a formulation it takes the samemeaning as the composition of the invention. Accordingly, formulationand composition are used interchangeably.

Throughout the description and claims of this specification, the words“comprise” and “contain” and variations of them mean “including but notlimited to”, and they are not intended to (and do not) exclude othermoieties, additives, components, integers or steps. Throughout thedescription and claims of this specification, the singular encompassesthe plural unless the context otherwise requires. In particular, wherethe indefinite article is used, the specification is to be understood ascontemplating plurality as well as singularity, unless the contextrequires otherwise.

Features, integers, characteristics, compounds, chemical moieties orgroups described in conjunction with a particular aspect, embodiment orexample of the invention are to be understood to be applicable to anyother aspect, embodiment or example described herein unless incompatibletherewith. All of the features disclosed in this specification(including any accompanying claims, abstract and drawings), and/or allof the steps of any method or process so disclosed, may be combined inany combination, except combinations where at least some of suchfeatures and/or steps are mutually exclusive. The invention is notrestricted to the details of any foregoing embodiments. The inventionextends to any novel one, or any novel combination, of the featuresdisclosed in this specification (including any accompanying claims,abstract and drawings), or to any novel one, or any novel combination,of the steps of any method or process so disclosed.

The reader's attention is directed to all papers and documents which arefiled concurrently with or previous to this specification in connectionwith this application and which are open to public inspection with thisspecification, and the contents of all such papers and documents areincorporated herein by reference.

Composition

The liquid composition comprises cyclosporin, a polymer capable offorming a matrix (a hydrogel forming polymer), a surfactant and an oilphase. The oil phase, cyclosporin and the surfactant are containedwithin the polymer capable of creating a matrix. The cyclosporin isdissolved in the oil phase. When the polymer forms a matrix the liquidcomposition is formed into a composition of the invention.

The composition comprises a matrix and cyclosporin. The matrix may beformed with a hydrogel-forming polymer, and may contain additionalexcipient(s) to the polymer. The active ingredient is contained withinthe matrix. The active ingredient may be in solution or in suspension,or in a combination thereof; however the invention is not limited tocompositions comprising a solution or suspension of the active and itincludes, for example, active ingredients encapsulated in liposomes orcyclodextrin. The matrix may contain inclusions in which the activeingredient is comprised; for example, the inclusions may comprise ahydrophobic medium in which the active ingredient is dissolved orsuspended. An active ingredient may therefore be directly dissolved orsuspended in the matrix, or it may be dissolved or suspended indirectlyin the matrix by way of inclusions in which the active ingredient isdissolved or suspended.

The composition, therefore, comprises a matrix-forming polymer, inparticular a hydrogel-forming polymer. The matrix of the composition maybe or comprise a polymer matrix comprising a polymer selected from awater-permeable polymer, a water-swellable polymer and a biodegradablepolymer. In particular, the matrix is or comprises a hydrogel-formingpolymer described in more detail below.

Modified release of the active ingredient from the composition may beachieved by virtue of the properties of the matrix material. For examplethe matrix may be a permeable or erodible polymer within which theactive ingredient is contained, e.g. dissolved or suspended; followingoral administration the matrix is gradually dissolved or eroded therebyreleasing the active ingredient from the matrix. Erosion may be achievedby biodegradation of a biodegradable polymer matrix. Where the matrix ispermeable, water permeates the matrix enabling the drug to diffuse fromthe matrix. A matrix formed with a hydrogel-forming polymer maytherefore include a modified release polymer. As such modified releasepolymers may be mentioned cellulose derivatives, for examplehydroxypropylmethyl cellulose, poly(lactic acid), poly(glycoloic)acid,poly(lactic-co glycolic acid copolymers), polyethylene glycol blockco-polymers, polyorthoesters, polyanhydrides, polyanhydride esters,polyanhydride imides, polyamides and polyphosphazines.

Water Soluble Cellulose Ether Coating

The invention provides pharmaceutical compositions that may have a firstcoating which is or comprises a water-soluble cellulose ether. Theinvention provides pharmaceutical compositions that have a first polymercoating, wherein the polymer is or comprises a water-soluble celluloseether. The water-soluble cellulose ether may be, for example selectedfrom methyl cellulose, hydroxyethyl cellulose, hydroxylpropyl celluloseand hydroxypropylmethyl cellulose.

Suitably the material of the first coating (i.e. the sub-coating) isdifferent to the second coating on the composition. For example, wherethe first coating is or comprises a water-soluble ester of a celluloseether, the major component(s) (e.g. more than 50%) of the second coatingis or comprises a different polymer to that of the first coating.Accordingly, the first and second coatings suitably provide two layersof material as part of the composition. It is to be understood that whenthe second coating comprises a mixture of components, minor componentsof the outer second coating may the same as the material of the firstcoating. By way of example, when the first coating is or comprises HPMCand the second coating comprises ethyl cellulose, the ethyl cellulosemay optionally further comprise a minor amount (e.g. less than 50%, 40%,30% or 20%) of the first coating material, HPMC in this example. In suchembodiments the sub-coat and the second coating are considered to bedifferent.

The water-soluble cellulose ether may be a water-soluble cellulose etherselected from an alkyl cellulose, for example methyl cellulose, ethylmethyl cellulose; a hydroxyalkyl cellulose, for example hydroxyethylcellulose (available as Cellosize™ and Natrosol™), hydroxypropylcellulose (available as Klucel™) or hydroxymethyl cellulose; ahydroxyalkyl alkyl cellulose, for example hydroxyethyl methyl cellulose(NEMC), hydroxypropyl methyl cellulose (available as Methocel™Pharmacoat™, Benecel™) or ethyl hydroxyethyl cellulose (EHEC); and acarboxyalkyl cellulose, for example carboxymethyl cellulose (CMC).Suitably the water-soluble cellulose ether may, for example be selectedfrom methyl cellulose, hydroxyethyl cellulose, hydroxylpropyl celluloseand hydroxypopylmethyl cellulose.

The water-soluble cellulose ether may be a low viscosity polymer whichis suitable for application as a film or coating to the composition. Theviscosity of the polymer may be from about 2 to about 60 mPa·s, forexample a viscosity of: about 2 to about 20 mPa·s; about to 2 to about 8mPa·s; more suitably a viscosity of about 4 to about 10 mPa·s, forexample about 4 to about 6 mPa·s. Alternatively, the viscosity of thepolymer may fall outside any or all of the just-mentioned ranges, forexample be above 20 mPa·s. Alternatively, the viscosity of the polymermay fall outside any or all of the just-mentioned ranges, for example beabove 20 mPa·s. The viscosity of the polymer may be determined bymeasuring the viscosity of a 2% solution of the polymer in water at 20°C. using a Ubbelode viscometer using ASTM standard methods (D1347 andD2363).

The water soluble cellulose ether may be a water-solublehydroxypopylmethyl cellulose (HPMC or hypromellose). HPMC is prepared bymodifying cellulose to substitute hydroxy groups with methoxy andhydroxypropyl groups. Each anhydroglucose unit in the cellulose chainhas three hydroxyl groups. The amount of substituent groups on theanhydroglucose units may be expressed as the degree of substitution. Ifall three hydroxyl groups on each unit are substituted, the degree ofsubstitution is 3. The number of substituent groups on the ringdetermines the properties of the HPMC. The degree of substitution mayalso be expressed as the weight of the methoxy and hydroxypropyl groupspresent. Suitably the HPMC has from about 19 to about 30% methoxysubstitution and from about 7 to about 12% hydroxypropyl substitution,Particularly the HPMC has 25 to 30% methoxy substitution and 7 to 12%hydroxypropyl substitution. Suitably the HPMC is a low viscosity HPMCwhich is suitable for application as a film or coating to thecomposition. The viscosity of the HPMC is suitably from about 2 to 60mPa·s, for example about 2 to about 20 mPa·s, more suitably a viscosityof about 4 to about 10 mPa·s. The viscosity of the HPMC is determined bymeasuring the viscosity of a 2% solution of the HPMC in water at 20° C.using a Ubbelode viscometer using ASTM standard methods (D1347 andD2363). Such HPMC is available as for example Methocel™, for exampleMethocel™ E, including Methocel™ E5.

When the first coating is or comprises a water-soluble derivative of acellulose ether, the derivative may, for example be a water-solubleester of a cellulose ether. Water-soluble esters of cellulose ethers arewell known and may comprise esters of a cellulose ether, formed with oneor more suitable acylating agent(s). Acylation agents may be, forexample suitable acids or acid anhydrides or acyl halides. Accordinglythe ester of a cellulose ether may contain a single ester moiety or twoor more ester moieties to give a mixed ester. Examples of water-solubleesters of cellulose ethers may be water-soluble phthalate, acetate,succinate, propionate or butyrate esters of a cellulose ether (forexample HPMC). Suitably the water-soluble ester of a cellulose ether isa water-soluble phthalate, acetate-succinate, propionate,acetate-propionate or acetate-butyrate ester of a cellulose ether (forexample HPMC).

In one embodiment the water-soluble ester of a cellulose ether may be orcomprise a water-soluble ester of any of the water-soluble celluloseethers described above in relation to the sub-coating.

Particular water-soluble esters of cellulose ethers are water-solubleesters of HPMC. Esters of HPMC which are soluble in water at a pHgreater than 5.5 may be or comprise hydroxypropyl methylcellulosephthalate (HPMCP), or hydroxypropyl methylcellulose acetate succinate(HPMCAS) in which the presence of ionisable carboxyl groups causes thepolymer to solubilize at high pH (>5.5 for the LF grade and >6.8 for theHF grade). These polymers are commercially available from Shin-EtsuChemical Co. Ltd.

The cellulose ether-containing coating may comprise or be hypromellose,e.g. it may be made of a mixture of hypromellose, titanium dioxide andpolyethylene glycol; the coating may comprise at least 20 wt %hypromellose and optionally at least 50% or at least 75 wt %hypromellose, e.g. at least 80 wt % or at least 85 wt % or 90 wt %hypromellose. The coating material used to form the coating maytherefore comprise a dry weight percentage of hypromellose mentioned inthe preceding sentence.

If it is desired for the coating to use a mixture of hypromellose,titanium dioxide and polyethylene glycol, commercial productscorresponding to such mixtures are available including Opadry White, aproduct commercialised by Colorcon. More generally, there may bementioned various products commercialised under the trade name Opadryand Opadry II. Further non limiting examples include Opadry YS-1-7706-Gwhite, Opadry Yellow 03692357, Opadry Blue 03690842). These formulationsare available as dry film coating formulations that can be diluted inwater shortly before use. Opadry and Opadry II formulations comprise acellulosic film forming polymer (e.g., HPMC and/or HPC), and may containpolydextrose, maltodextrin, a plasticizer (e.g., triacetin, polyethyleneglycol), polysorbate 80, a colorant (e.g., titanium dioxide, one or moredyes or lakes), and/or other suitable film-forming polymers (e.g.,acrylate-methacrylate copolymers). Suitable OPADRY or OPADRY IIformulations may comprise a plasticizer and one or more of maltodextrin,and polydextrose (including but not limited to a) triacetin andpolydextrose or maltodextrin or lactose, or b) polyethylene glycol andpolydextrose or maltodextrin). Particularly preferred commercialproducts are Opadry White (HPMC/HPC-based) and Opadry II White(PVA/PEG-based).

The cellulose ether-containing coating may also be applied as a simplesolution comprising water and the polymer of the first coating. Forexample when the polymer is an HPMC, for example such as Methocel, thefirst coating may be applied to the core as an aqueous solution ordispersion of the HPMC. Optionally the coating solution may includeother solvents such as an alcohol. Alternatively the coating may beapplied as a solution or dispersion in a volatile organic solvent.

Suitably the first coating that contains a water soluble cellulose etheris present in an amount corresponding to a weight gain of thecomposition due to the coating of from 0.5% to 40% (for example from0.5% to 30%; from 0.5% to 20%; from 1% to 25%; from 1% to 15%; from 1%to 6%; from 1% to 4%; from 4% to 6%; from 6% to 10%; from 9% to 15%; orfrom 12% to 15%) by weight based upon the weight of the compositionprior to applying the coating. The first coating that contains a watersoluble cellulose ether is present in an amount corresponding to aweight gain of the composition due to the coating of from 1% to 10%;from 4% to 10%; from 4% to 8%; and from 5% to 8% by weight based uponthe weight of the core or the composition prior to applying the coating.

In another embodiment the first coating that contains a water-solublecellulose ether is present in an amount corresponding to a weight gaindue to the first coating in a range selected from 9 to 30%, suitably 9%to 20%, or particularly 10% to 15% by weight based upon the weight ofthe composition prior to applying the coating.

Suitably the first coating that contains a water soluble cellulose etherprovides a coating thickness on the composition of at least 5 μm,suitably from about 5 μm to about 1 mm, for example from about 10 μm toabout 1 mm, from about 10 μm to about 500 μm, from about 50 μm to about1 mm, or about from about 50 μm to about 500 μm. The thickness maytherefore be from about 100 μm to about 1 mm, e.g. 100 μm to about 750μm or about 100 μm to about 500 μm. The thickness may be from about 250μm to about 1 mm, e.g. about 250 μm to about 750 μm or 250 μm to about500 μm. The thickness may be from about 500 μm to about 1 mm, e.g. about750 μm to about 1 mm or about 500 μm to about 750 μm. The thickness maytherefore be from about 10 μm to about 100 μm, e.g. from about 10 μm toabout 50 μm or about 50 μm to about 100 μm.

When the first coating comprises a water-soluble cellulose ether thecellulose ether(s) suitably forms at least 40%, 50%, 60%, 70%, 80%, 85%or 90% by weight of the dry weight of the first coating. Alternativelythe water-soluble cellulose ether is the first coating.

It is preferred to dry the composition of the invention before the firstcoating that contains a water-soluble cellulose ether is applied, as isdescribed in more detail below in relation to the coating process.

It has been found that applying to a core comprising a pharmaceuticallyactive ingredient a sub-coating, referred to elsewhere in theapplication as the subcoat (hence the subcoat and the first coating areequivalent), that contains a water soluble cellulose ether prior toapplying a delayed release coating provides unexpected advantages. Thepresence of such a sub-coating has been found to enhance the dissolutionproperties of the delayed release compositions according to theinvention. In particular the presence of such a sub-coating has beenfound to increase the rate of release of the active ingredient from thecomposition and also to increase the amount of the active ingredientreleased in a set time period compared to compositions prepared withoutusing such a sub-coating. These findings are unexpected, because itwould have been expected that the presence of a sub-coating in additionto a delayed release outer coating would act to delay or inhibit releaseof drug from the composition and, at a given time, for there to be lessdrug released, because there is a thicker coating present. However, asillustrated in the Examples, contrary to these expectations both theextent and rate of release of active ingredient are increased comparedto compositions without such a sub-coating. Accordingly, delayed releasecompositions according to the invention which comprise a sub-coat thatcomprises or is a water-soluble cellulose ether and a delayed releasecoating outside the sub-coat, provide a unique dissolution profile. Thepresence of such a sub-coating has also been found to reducebatch-to-batch variability, particularly when the core is in the form ofa minibead. A sub-coating that comprises or is a water-soluble celluloseether may therefore also reduce intra- and inter-patient variability asa result of a more consistent dissolution profile. The unique propertiesof sub-coated compositions according to the invention (particularly thedissolution profile) are expected to contribute to favourablepharmacokinetic properties of the compositions according to theinvention.

Accordingly in an embodiment there is provided a composition comprisingcyclosporin, a hydrogel forming polymer matrix, a surfactant and an oilphase being dispersed in the hydrogel forming polymer matrix, whereinthe surfactant is a medium chain or long chain fatty acid mono- ordi-glyceride or a combination thereof and does not comprise or is not apolyethyleneglycol ether or ester, the composition further comprising afirst coating; and wherein

the first coating is or comprises a water-soluble cellulose ether.

The composition may have a second coating comprising or being a delayedrelease polymer.

Accordingly in an embodiment there is provided a composition comprisingcyclosporin, a hydrogel forming polymer matrix, a surfactant and an oilphase being dispersed in the hydrogel forming polymer matrix, whereinthe surfactant is a medium chain or long chain fatty acid mono- ordi-glyceride or a combination thereof and does not comprise or is not apolyethyleneglycol ether or ester, the composition further comprising afirst coating and a second coating outside the first coating; andwherein

the first coating is or comprises a water-soluble cellulose ether; and

the second coating is or comprises a delayed release coating, e.g. is orcomprises a delayed release polymer.

An aspect of the invention resides in a multiple minibead compositioncomprising at least two populations of active ingredient-containingminibeads, wherein members of at least one minibead population areminibeads as described herein (i.e. compositions of the invention inminibead format). It will be understood that the two populations aredifferent. Such a plural minibead population composition may comprise orconsist of the following two populations:

-   -   a first population having a coating that is or comprises a        water-soluble cellulose ether but having no outer coating, e.g.        as described herein; and    -   a second population having a first coating that is or comprises        a water-soluble cellulose ether and a second coating that is or        comprises a delayed release coating, for example as described        herein e.g. a coating that is or comprises a delayed release        polymer.

The respective minibeads of each population of a plural minibeadcomposition may contain cyclosporin as the minibeads of some or all ofthe other populations, or one population may contain cyclosporin andanother population may contain a different active ingredient(s) thereto,e.g. a different combination.

A multiple population composition may be for use in treating a disorderof the GI tract, for example as described herein. Such a composition maybe for use in treating a disorder affecting multiple regions of the GIT,e.g. the upper intestine and the lower intestine, and may comprise anactive ingredient selected from immunosuppressants (e.g. cyclosporin),hydroxylase inhibitors (e.g. hydralazine) and anti-inflammatories (e.g.mesalazine).

The minibeads of a multiple population composition may by way of examplebe contained in a gel capsule or a sachet.

The second coating is outside the first coating and may be any of thedelayed release coatings described herein. In particular, the secondcoating is or comprises a pH independent polymer modified releasecoating described above. For example the second coating may be orcomprise an enteric coating or a pH independent coating. The secondcoating may comprise a mixture of polymers including a polymerdegradable by bacterial or other enzymes. In a particular embodiment thesecond coating comprises ethyl cellulose and optionally a water-solublepolysaccharide, in particular one susceptible to degradation by colonicbacteria, suitably pectin. Accordingly the second coating may comprisethe Surelease-pectin mixture described above.

It is not a requirement that both the first and second coatings arepresent in the composition at the same time. For example, thecomposition may comprise second coating (outer coating) in the absenceof a first coating. Conversely, the composition may comprise a firstcoating in the absence of a second coating.

The first and second coating may independently be aqueous-based coatingsor may be solvent-based coatings. By this it is meant that the firstand/or second coating may be formulated prior to being applied to thecore or composition and/or applied to the core or composition as anaqueous-based composition or as a solvent-based (non-aqueoussolvent-based) composition. The aqueous-based or solvent-based coatingcompositions may be a suspension or a solution of the coating materialin water or in a solvent.

In an embodiment the composition comprises a core and an outer coating(also referred to as a second coating herein), the core comprisingcyclosporin, a hydrogel forming polymer matrix, a surfactant and an oilphase being dispersed in the hydrogel forming polymer matrix, whereinthe surfactant is a medium chain or long chain fatty acid mono- ordi-glyceride or a combination thereof and does not comprise or is not apolyethyleneglycol ether or ester. The composition may optionallyfurther comprise a sub-coat.

In one embodiment of the invention there is provided a compositioncomprising a core, a first coating and a second coating outside thefirst coating; and wherein:

the core comprises cyclosporin, a hydrogel forming polymer matrix, asurfactant and an oil phase being dispersed in the hydrogel formingpolymer matrix, wherein the surfactant is a medium chain or long chainfatty acid mono- or di-glyceride or a combination thereof and does notcomprise or is not a polyethyleneglycol ether or ester;

the first coating is or comprises a water-soluble cellulose ether,particularly hydroxypropylmethyl cellulose;

the second coating is or comprises a modified release coating or delayedrelease coating, particularly a pH independent modified release coating;

the first coating is present in an amount corresponding to a weight gaindue to the first coating in a range selected from: (i) 8% to 15%; (ii)from 8% to 12%, for example about 10%; or (iii) from 2.5% to 6%, forexample about 5% by weight based upon the weight of the compositionprior to applying the first coating; and wherein

the second coating is present in an amount corresponding to a weightgain of the composition due to the second coating selected from (a) from4% to 20%; (b) from 7.5% to 20%; (C) from 10% to 12%, for example about11% or about 11.5%; or (d) from 16% to 18%, for example about 17% byweight based upon the weight of the composition prior to applying thesecond coating.

The first and second coatings in the embodiment of the immediatelypreceding paragraph are suitably any of the first and second coatingsdescribed above or below. Accordingly it is intended that the coatingsdescribed in this section may be applied to any of the compositionsdescribed herein to provide a delayed release coating if required. Thecoatings are particularly useful to provide a modified release coatingto the cores comprising a polymer matrix and pharmaceutically activeingredient described in this application.

Outer Barrier or Protective Coating

The compositions described herein may comprise a protective coatingoutside the first and/or second coating, for example outside the secondcoating, the modified release coating. The protective coating may helpto protect the modified release coating from damage resulting from, forexample formulating the composition into a final dosage form, or duringthe handling, transport or storage of the composition. The protectivecoating is suitably applied to the outer surface of the composition. Theprotective coating may be applied directly to the second coating (themodified release coating) such that the protective coating is in contactwith the second coating (the modified release coating). The protectivecoating is suitably a water soluble coating which does not adverselyaffect the release of the cyclosporin A from the composition when inuse. Suitably the protective coating is or comprises a water-solublepolymer. The protective coating may comprise a water-soluble cellulosicor PVA film-forming polymer. Suitably the protective coating may be orcomprise Opadry (HPMC/HPC-based), Opadry II (PVA/PEG-based) or polyvinylalcohol-polyethylene glycol graft copolymers (Kollicoat IR) as describedherein. The protective coating may be present as a layer of from about 2to about 50 μm. Suitably the protective coating is applied to give aweight-gain of from about 0.5 to about 10%, based upon the weight of thecomposition prior to applying the protective coating.

Polymer Matrix

The composition of the invention comprises cyclosporin, a hydrogelforming polymer matrix, a surfactant and an oil phase being dispersed inthe hydrogel forming polymer matrix. In addition, in certain embodimentsof the invention the composition of the invention comprises a corewherein the core comprises cyclosporin, a hydrogel forming polymermatrix, a surfactant and an oil phase being dispersed in the hydrogelforming polymer matrix. The composition or the core comprises acontinuous phase or matrix phase, which may be or comprise the hydrogelforming polymer matrix, to provide mechanical strength. In embodimentsthe cyclosporin is comprised within a disperse phase or oil phase withinthe continuous phase or matrix. The cyclosporin may be present as adisperse phase within the hydrogel-forming polymer matrix (continuousphase or aqueous phase) of the core or composition. The disperse phasemay be or comprise the oil phase. For example the disperse phase maycomprise a lipid and cyclosporin A. The core or the composition may beprepared by dispersing the cyclosporin, dissolved in the oil phasewithin an aqueous phase comprising the hydrogel forming polymer matrixto form a colloid and then causing the composition to solidify (gel),thereby immobilising the cyclosporin within the hydrogel-forming polymermatrix.

The core may have the form of a solid colloid, the colloid comprising acontinuous phase and a disperse phase, wherein the continuous phase isor comprises the hydrogel-forming polymer matrix and the disperse phaseis or comprises an oil phase optionally comprising the cyclosporin. Thedisperse phase may comprise a vehicle containing the cyclosporin, forexample containing it as a solution or a suspension or a combination ofboth. The vehicle may be an oil phase as described herein.

Such cores comprising a hydrogel-forming polymer and a disperse phasecomprising cyclosporin A are described in more detail below.

Delayed Release Coatings

The invention provides compositions having a coating that comprises, oris, a coating-forming polymer, wherein the coating-forming polymer is ahydrogel-forming polymer; the coating may be a first coating outsidewhich is a second coating. The second coating may be a delayed releasecoating, although the invention does not require that the second coatingbe a delayed release coating. The second coating may comprise or be adelayed release polymer.

The first coating may be present in an amount described elsewhere inthis specification.

The first coating may be present in an amount corresponding to a weightgain due to the first coating of from 0.5% to 20% by weight of the core.

Furthermore, the composition may comprise a first coating present in anamount corresponding to a weight gain due to the coating selected fromranges of from: 0.5% to 15%; 1% to 15%; 1% to 12%; 1% to 10%; 1% to 8%;1% to 6%; 1% to 4%, 2% to 10%; 2% to 8%; 2% to 6%; 2% to 7%; 2% to 4%;4% to 8%; 4% to 7%, 4% to 6%, 5% to 7%; 7% to 20%; 7% to 16%; 9% to 20%;9% to 16%; 10% to 15%; and 12% to 16%.

The invention provides for a pharmaceutical composition comprising acore, a first coating and a second coating outside of the first coating,wherein the core comprises cyclosporin, a hydrogel forming polymermatrix, a surfactant and an oil phase being dispersed in the hydrogelforming polymer matrix, the first coating comprises or is a watersoluble cellulose ether, and the second coating comprises or is adelayed release polymer, and the first coating may be present in anamount corresponding to a weight gain due to the first coating of from0.5% to 20% by weight of the core, wherein the surfactant is a mediumchain or long chain fatty acid mono- or di-glyceride or a combinationthereof and does not comprise or is not a polyethyleneglycol ether orester.

The composition of the invention may comprise a first coating with athickness of 1 μm to 1 mm. Thus, the % weight gain due to the coatingspecified above may correspond to a thickness of 1 μm to 1 mm.

The first coating may have a thickness selected from ranges of from: 1μm to 500 μm; 10 μm to 250 μm; 10 μm to 100 μm; 10 μm to 50 μm; 10 μm to20 μm; 50 μm to 100 μm; 100 μm to 250 μm; 100 μm to 500 μm; 50 μm to 500μm; 50 μm to 250 μm; 100 μm to 1 mm; 500 μm to 1 mm. The coating havingthe thicknesses disclosed in this paragraph may be any of the coatingsin the application. In particular the coating referred to in thisparagraph may be the water-soluble cellulose ether coating.

The first coating may be present in a weight gain selected from a rangeof from: 1% to 20%, 4% to 7%, 5% to 7%, 4% to 15%, 8% to 15%, 4% to 12%and 8% to 12%. The second coating may be present in a weight gainselected from a range of from: 8% to 15% or 8% to 12%.

In addition, the invention provides for a pharmaceutical compositioncomprising a core, a first coating and a second coating outside of thefirst coating, wherein the core comprises cyclosporin, a hydrogelforming polymer matrix, a surfactant and an oil phase being dispersed inthe hydrogel forming polymer matrix, the first coating comprises or is awater soluble cellulose ether, and the second coating comprises or is adelayed release polymer, and the first coating has a thickness of from 1μm to 1 mm. The core may optionally further comprise a hydrogel formingpolymer, wherein the surfactant is a medium chain or long chain fattyacid mono- or di-glyceride or a combination thereof and does notcomprise or is not a polyethyleneglycol ether or ester.

The second coating may be present in an amount described elsewhereherein. Suitably the second coating provides a coating thickness on thecomposition of from about 10 μm to about 1 mm, for example, from about10 μm to about 500 μm, from about 50 μm to about 1 mm, or about fromabout 50 μm to about 500 μm. The thickness may therefore be from about100 μm to about 1 mm, e.g. 100 μm to about 750 μm or about 100 μm toabout 500 μm. The thickness may be from about 250 μm to about 1 mm, e.g.about 250 μm to about 750 μm or 250 μm to about 500 μm. The thicknessmay be from about 500 μm to about 1 mm, e.g. about 750 μm to about 1 mmor about 500 μm to about 750 μm. The thickness may therefore be fromabout 10 μm to about 100 μm, e.g. from about 10 μm to about 50 μm orabout 50 μm to about 100 μm.

It is contemplated within any aspect or embodiment where there is asecond coating (also referred to as an outer coating) that the secondcoating may be present in a % weight gain of from 2% to 40%. In additionthe second coating may be present in an amount corresponding to a weightgain due to the coating selected from ranges of from: 4% to 30%, 4% to7%, 7% to 40%, 7% to 30%, 8% to 25%, 8% to 20%, 2% to 25%, 2% to 20%, 4%to 25%, 4% to 20%, 4% to 15%, 4% to 13%, 7% to 15%, 7% to 13%, 8% to12%, 9% to 12% and 20% to 25%.

In any aspect and embodiment of the invention the first coating may bepresent in a % weight gain relative to the core of from 0.5% to 20%,preferably 1% to 16% or 4% to 16%, and the second coating may be presentin a % weight gain of 4% to 24%, 7% to 24%, 22% to 24%, 7% to 15%, or 8%to 12%, preferably 22% to 24%, 7% to 15%, or 8% to 12%.

The core is preferably in the form of a minibead, for example asdescribed hereafter in more detail, for example in the form of a solidcolloid. The second coat may be a film or it may be a membrane. Thesecond coat, e.g. film or membrane, may serve to delay release untilafter the stomach; the coat may therefore be an enteric coat. Thedelayed release coat may comprise one or more delayed releasesubstances, preferably of a polymeric nature (e.g. methacrylates etc;polysaccharides etc as described in more detail below), or combinationof more than one such substance, optionally including other excipients,for example, plasticizers. Preferred plasticizers, if they are used,include hydrophilic plasticizers for example triethyl citrate (TEC)which is particularly preferred when using the Eudragit® family ofpolymers as coatings as described below. Another preferred plasticiser,described in more detail below in relation to coating with ethylcellulose, is dibutyl sebacate (DBS). Alternative or additionaloptionally included excipients are glidants. A glidant is a substancethat is added to a powder or other medium to improve its flowability. Atypical glidant is talc which is preferred when using the Eudragit®family of polymers as coatings.

The delayed release coating (the second coating) may be applied asdescribed below and may vary as to thickness and density. The amount ofcoat is defined by the additional weight added to (gained by) the drycomposition (e.g. the core) to which it is applied. Weight gain ispreferably in the range 0.1% to 50%, preferably from 1% to 15% of thedry weight of the core, more preferably in the range 3% to 10% or in therange 5-12% or in the range 8-12%.

Polymeric coating material of a delayed release coating may comprisemethacrylic acid co-polymers, ammonio methacrylate co-polymers, ormixtures thereof. Methacrylic acid co-polymers such as, for example,EUDRAGIT™ S and EUDRAGIT™ L (Evonik) are particularly suitable. Thesepolymers are gastroresistant and enterosoluble polymers. Their polymerfilms are insoluble in pure water and diluted acids. They may dissolveat higher pHs, depending on their content of carboxylic acid. EUDRAGIT™S and EUDRAGIT™ L can be used as single components in the polymercoating or in combination in any ratio. By using a combination of thepolymers, the polymeric material can exhibit solubility at a variety ofpH levels, e.g. between the pHs at which EUDRAGIT™ L and EUDRAGIT™ S areseparately soluble. In particular, the coating may be an enteric coatingcomprising one or more co-polymers described in this paragraph. Aparticular coating material to be mentioned is Eudragit L 30 D-55.

The trade mark “EUDRAGIT” is used hereinafter to refer to methacrylicacid copolymers, in particular those sold under the trade mark EUDRAGITby Evonik.

The delayed release coating, where present, can comprise a polymericmaterial comprising a major proportion (e.g., greater than 50% of thetotal polymeric coating content) of at least one pharmaceuticallyacceptable water-soluble polymer, and optionally a minor proportion(e.g., less than 50% of the total polymeric content) of at least onepharmaceutically acceptable water insoluble polymer. Alternatively, themembrane coating can comprise a polymeric material comprising a majorproportion (e.g., greater than 50% of the total polymeric content) of atleast one pharmaceutically acceptable water insoluble polymer, andoptionally a minor proportion (e.g., less than 50% of the totalpolymeric content) of at least one pharmaceutically acceptablewater-soluble polymer.

Ammonio methacrylate co-polymers such as, for example, EUDRAGIT™ RS andEUDRAGIT™ RL (Evonik) are suitable for use in the present invention.These polymers are insoluble in pure water, dilute acids, buffersolutions, and/or digestive fluids over the entire physiological pHrange. The polymers swell in water and digestive fluids independently ofpH. In the swollen state, they are then permeable to water and dissolvedactive agents. The permeability of the polymers depends on the ratio ofethylacrylate (EA), methyl methacrylate (MMA), and trimethylammonioethylmethacrylate chloride (TAMCl) groups in the polymer. For example, thosepolymers having EA:MMA:TAMCl ratios of 1:2:0.2 (EUDRAGIT™ RL) are morepermeable than those with ratios of 1:2:0.1 (EUDRAGIT™ RS). Polymers ofEUDRAGIT™ RL are insoluble polymers of high permeability. Polymers ofEUDRAGIT™ RS are insoluble films of low permeability. Adiffusion-controlled pH-independent polymer in this family is RS 30 Dwhich is a copolymer of ethyl acrylate, methyl methacrylate and a lowcontent of methacrylic acid ester with quaternary ammonium groupspresent as salts to make the polymer permeable. RS 30 D is available asan aqueous dispersion.

The amino methacrylate co-polymers can be combined in any desired ratio,and the ratio can be modified to modify the rate of drug release. Forexample, a ratio of EUDRAGIT™ RS:EUDRAGIT™ RL of 90:10 can be used.Alternatively, the ratio of EUDRAGIT™ RS:EUDRAGIT™ RL can be about 100:0to about 80:20, or about 100:0 to about 90:10, or any ratio in between.In such compositions, the less permeable polymer EUDRAGIT™ RS generallycomprises the majority of the polymeric material with the more solubleRL, when it dissolves, permitting gaps to be formed through whichsolutes can come into contact with the core allowing for the active toescape in a controlled manner.

The amino methacrylate co-polymers can be combined with the methacrylicacid co-polymers within the polymeric material in order to achieve thedesired delay in the release of the drug and/or poration of the coatingand/or exposure of the composition within the coating to allow egress ofdrug and/or dissolution of the immobilization or water-soluble polymermatrix. Ratios of ammonio methacrylate co-polymer (e.g., EUDRAGIT™ RS)to methacrylic acid co-polymer in the range of about 99:1 to about 20:80can be used. The two types of polymers can also be combined into thesame polymeric material, or provided as separate coats that are appliedto the beads.

Eudragit™ FS 30 D is an anionic aqueous-based acrylic polymericdispersion consisting of methacrylic acid, methyl acrylate, and methylmethacrylate and is pH sensitive. This polymer contains fewer carboxylgroups and thus dissolves at a higher pH (>6.5). The advantage of such asystem is that it can be easily manufactured on a large scale in areasonable processing time using conventional powder layering andfluidized bed coating techniques. A further example is EUDRAGIT® L30D-55 which is an aqueous dispersion of anionic polymers withmethacrylic acid as a functional group. It is available as a 30% aqueousdispersion.

In addition to the EUDRAGIT™ polymers described above, a number of othersuch copolymers can be used to control drug release. These includemethacrylate ester co-polymers such as, for example, the EUDRAGIT™ NEand EUDRAGIT™ NM ranges. Further information on the EUDRAGIT™ polymerscan be found in “Chemistry and Application Properties ofPolymethacrylate Coating Systems,” in Aqueous Polymeric Coatings forPharmaceutical Dosage Forms, ed. James McGinity, Marcel Dekker Inc., NewYork, pg 109-114 the entirety of which is incorporated herein byreference.

Several derivatives of hydroxypropyl methylcellulose (HPMC) also exhibitpH dependent solubility and may be used in the invention for the delayedrelease coating. As examples of such derivatives may be mentioned HPMCesters, for example hydroxypropyl methylcellulose phthalate (HPMCP),which rapidly dissolves in the upper intestinal tract and hydroxypropylmethylcellulose acetate succinate (HPMCAS) in which the presence ofionisable carboxyl groups causes the polymer to solubilize at high pH(>5.5 for the LF grade and >6.8 for the HF grade). These polymers arecommercially available from Shin-Etsu Chemical Co. Ltd. As with otherpolymers described herein as useful for delayed release coatings, HPMCand derivatives (e.g. esters) may be combined with other polymers e.g.EUDRAGIT RL-30 D.

Other polymers may be used to provide a coating in particular enteric,or pH-dependent, polymers. Such polymers can include phthalate,butyrate, succinate, and/or mellitate groups. Such polymers include, butare not limited to, cellulose acetate phthalate, cellulose acetatesuccinate, cellulose hydrogen phthalate, cellulose acetate trimellitate,hydroxypropyl-methylcellulose phthalate, hydroxypropylmethylcelluloseacetate succinate, starch acetate phthalate, amylose acetate phthalate,polyvinyl acetate phthalate, and polyvinyl butyrate phthalate.

pH Independent Polymer Delayed Release Coatings

In a particular embodiment the second coating, where present, is orcomprises a polymeric coating which is pH-independent in its dissolutionprofile and/or in its ability to release the active ingredientincorporated in the compositions of the invention. A pH-independentpolymer delayed release coating comprises a delayed release polymer,optionally a plurality of delayed release polymers, and one or moreother optional components. The other components may serve to modulatethe properties of the composition. Examples have already been given(e.g., Eudragit RS and RL).

Another example of a pH-independent polymeric coating is a coating thatcomprises or is ethylcellulose; a pH-independent polymeric coating mayhave a delayed release polymer that is ethylcellulose, therefore. Itwill be understood that an ethylcellulose formulation for use in coatinga dosage form may comprise, in addition to ethylcellulose and—in thecase of a liquid formulation—a liquid vehicle, one or more othercomponents. The other components may serve to modulate the properties ofthe composition, e.g. stability or the physical properties of thecoating such as the flexibility of the film coating. The ethylcellulosemay be the sole delayed release polymer in such a composition. Theethylcellulose may be in an amount of at least 50%, at least 60%, atleast 70%, at least 80%, at least 90% or at least 95% by weight of thedry weight of a coating composition for use in coating a dosage form.Accordingly, an ethylcellulose coating may include other components inaddition to the ethylcellulose. The ethylcellulose may be in an amountof at least 50%, at least 60%, at least 70%, at least 80%, at least 90%or at least 95% by weight of the ethylcellulose coating. Consequently,ethylcellulose may be in an amount of at least 50%, at least 60%, atleast 70%, at least 80%, at least 90% or at least 95% by weight of thedry weight of the second coating. Suitably the ethyl cellulose coatingfurther comprises a plasticizer as described below to improve theflexibility of the film and to improve the film-forming properties ofthe coating composition during application of the coating.

A particular ethylcellulose coating composition which may be applied tothe composition, optionally to the core (i.e. in the absence of a firstcoating) or to the first coating is a dispersion of ethylcellulose in asub-micron to micron particle size range, e.g. from about 0.1 to 10 μmin size, homogeneously suspended in water with the aid of anemulsification agent, e.g. ammonium oleate. The ethylcellulosedispersion may optionally and preferably contain a plasticizer. Suitablyplasticisers include for example dibutyl sebacate (DBS),diethylphthalate, triethyl citrate, tributyl citrate, triacetin, ormedium chain triglycerides. The amount of plasticizer present in thecoating composition will vary depending upon the desired properties ofthe coating. Typically the plasticizer comprises from 1 to 50%, forexample about 8 to about 50% of the combined weight of the plasticizerand ethyl cellulose. Such ethylcellulose dispersions may, for example,be manufactured according to U.S. Pat. No. 4,502,888, which isincorporated herein by reference. One such ethylcellulose dispersionsuitable for use in the present invention and available commercially ismarketed under the trademark Surelease®, by Colorcon of West Point, Pa.USA. In this marketed product, the ethylcellulose particles are, e.g.,blended with oleic acid and a plasticizer, then optionally extruded andmelted. The molten plasticized ethylcellulose is then directlyemulsified, for example in ammoniated water optionally in a high shearmixing device, e.g. under pressure. Ammonium oleate can be formed insitu, for instance to stabilize and form the dispersion of plasticizedethylcellulose particles. Additional purified water can then be added toachieve the final solids content. See also U.S. Pat. No. 4,123,403,which is incorporated herein by reference.

The trademark “Surelease®” is used hereinafter to refer toethylcellulose coating materials, for example a dispersion ofethylcellulose in a sub-micron to micron particle size range, e.g. fromabout 0.1 to 10 μm in size, homogeneously suspended in water with theaid of an emulsification agent, e.g. ammonium oleate. In particular, thetrademark “Surelease®” is used herein to refer to the product marketedby Colorcon under the Surelease® trademark.

Surelease® dispersion is an example of a combination of film-formingpolymer, plasticizer and stabilizers which may be used as a secondcoating to adjust rates of active principle release with reproducibleprofiles that are relatively insensitive to pH. The principal means ofdrug release is by diffusion through the Surelease® dispersion membraneand is directly controlled by film thickness. Use of Surelease® isparticularly preferred and it is possible to increase or decrease thequantity of Surelease® applied as coating in order to modify thedissolution of the coated composition. Unless otherwise stipulated, useof the term “Surelease” may apply to Surelease E-7-19020, E-7-19030,E-7-19040 or E-7-19050. An ethylcellulose coating formulation, forexample Surelease E-7-19020, may comprise ethylcellulose blended witholeic acid and dibutyl sebacate, then extruded and melted. The moltenplasticized ethylcellulose is then directly emulsified in ammoniatedwater in a high shear mixing device under pressure. Ammonium oleate isformed in situ to stabilize and form the dispersion of plasticizedethylcellulose particles. Additional purified water is then added toachieve the final solids content. An ethylcellulose coating formulation,for example Surelease E-7-19030, may additionally comprise colloidalanhydrous silica dispersed into the material. An ethylcellulose coatingformulation, for example Surelease E-7-19040, may comprise medium chaintriglycerides instead of dibutyl sebacate, in particular in aformulation comprising colloidal anhydrous silica and oleic acid. Anethylcellulose coating formulation, for example Surelease E-7-19050, mayderive from blending ethylcellulose with oleic acid before melting andextrusion. The molten plasticized ethylcellulose is then directlyemulsified in ammoniated water in a high shear mixing device underpressure. Ammonium oleate is formed in situ to stabilize and form thedispersion of plasticized ethylcellulose particles. However,formulations that comprise medium chain triglycerides, colloidalanhydrous silica and oleic acid are preferred. Surelease E-7-19040 isparticularly preferred.

The invention also contemplates using combinations of ethylcellulose,e.g. a Surelease formulation, with other coating components, for examplesodium alginate, e.g. sodium alginate available under the trade nameNutrateric™.

In addition to the EUDRAGIT™ and Surelease® polymers discussed above,where compatible, any combination of coating polymers disclosed hereinmay be blended to provide additional delayed-release profiles.

The delayed release coating can further comprise at least one solubleexcipient to increase the permeability of the polymeric material. Thesesoluble excipients can also be referred to or are pore formers.Suitably, the at least one soluble excipient or pore former is selectedfrom among a soluble polymer, a surfactant, an alkali metal salt, anorganic acid, a sugar, a polysaccharide, and a sugar alcohol. Suchsoluble excipients include, but are not limited to, polyvinylpyrrolidone, polyvinyl alcohol (PVA), polyethylene glycol, awater-soluble hydroxypropyl methyl cellulose, sodium chloride,surfactants such as, for example, sodium lauryl sulfate andpolysorbates, organic acids such as, for example, acetic acid, adipicacid, citric acid, fumaric acid, glutaric acid, malic acid, succinicacid, and tartaric acid, sugars such as, for example, dextrose,fructose, glucose, lactose, and sucrose, sugar alcohols such as, forexample, lactitol, maltitol, mannitol, sorbitol, and xylitol, xanthangum, dextrins, and maltodextrins; and a polysaccharide susceptible ofdegradation by a bacterial enzyme normally found in the colon, forexample polysaccharides include chondroitin sulphate, pectin, dextran,guar gum and amylase, chitosan etc. and derivatives of any of theforegoing. In some embodiments, polyvinyl pyrrolidone, mannitol, and/orpolyethylene glycol can be used as soluble excipients. The at least onesoluble excipient can be used in an amount ranging from about 0.1% toabout 15% by weight, based on the total dry weight of the polymercoating, for example from about 0.5% to about 10%, about 0.5% to about5%, about 1% to about 3%, suitably about 2% based on the total dryweight of the polymer coating. The delayed release coating may be freefrom HPMC.

The modifications in the rates of release, such as to create a delay orextension in release, can be achieved in any number of ways. Mechanismscan be dependent or independent of local pH in the intestine, and canalso rely on local enzymatic activity to achieve the desired effect.Examples of modified-release compositions are known in the art and aredescribed, for example, in U.S. Pat. Nos. 3,845,770; 3,916,899;3,536,809; 3,598,123; 4,008,719; 5,674,533; 5,059,595; 5,591,767;5,120,548; 5,073,543; 5,639,476; 5,354,556; and 5,733,566 all of whichare incorporated herein by reference in their entirety.

The addition to Surelease® or other pH-independent polymer substance ofa second polymer (e.g. a polysaccharide, especially aheteropolysaccharide) which is susceptible to degradation by colonicbacterial enzymes (and optionally or alternatively by pancreatic orother relevant enzymes), helps provide targeted release of the activeingredient to a site or sites within the GI tract where the secondpolymer is degraded. By varying the amount of second polymer addedpresent in the coating the dissolution profile may be optimized toprovide the required release of cyclosporin A from the composition.

In a particular embodiments the delayed release coating provides forrelease of the active agent in at least the colon. Accordingly in oneembodiment the coating comprises a combination of ethylcellulose(preferably a described above, and particularly formulated with anemulsification agent such as, for example, ammonium oleate and/or aplasticizer such as, for example, dibutyl sebacate or medium chaintriglycerides) and a polysaccharide susceptible of degradation by abacterial enzyme normally found in the colon. Such polysaccharidesinclude chondroitin sulphate, pectin, dextran, guar gum and amylase,chitosan etc. and derivatives of any of the foregoing. Chitosan may beused in connection with obtaining a colon-specific release profile;additionally or alternatively, pectin may be so used.

The use of polysaccharides by themselves for delayed release coatingpurposes has been tried with limited success. Most of the non-starchpolysaccharides suffer from the drawback of lacking good film formingproperties. Also, they tend to swell in the GI tract and become porous,resulting in the early release of the drug. Even amorphous amylose,which is resistant to degradation by pancreatic alpha amylase butcapable of degradation by colonic bacterial enzymes, has thedisadvantage of swelling in aqueous media although this can becontrolled by incorporating insoluble polymer, for example ethylcellulose and/or acrylate, into the amylose film. Amylose however is notwater-soluble and although water-insoluble polysaccharides are notexcluded, use of a water-soluble polysaccharide (WSP) susceptible tobacterial enzymatic degradation brings particularly advantageous resultswhen used as a coating in accordance with this embodiment of the presentinvention. A particularly preferred polysaccharide in this embodiment ofthe present invention is pectin. Various kinds of pectin may be usedincluding pectin of different grades available i.e. with differingdegrees of methylation (DM), i.e. percentage of carbonyl groupsesterified with methanol, for example pectins with a DM of more than50%, known as High Methoxy (HM) Pectins or Low Methoxy (LM) pectins, ora pectin combination comprising an HM pectin and an LM pectin. It isalso possible in this embodiment to use pectins having various degreesof acetylation (Dac). Taken together, the DM and Dac or the degree ofsubstitution is known as Degree of Esterification (DE). Pectins ofvarious DE's may be used according to the invention. As an alternativeto pectin, sodium alginate may be used as a polysaccharide according toan embodiment of the invention. However, other embodiments mayconveniently include amylose and/or starch which contains amylose.Various grades of starch, containing different percentages of amylosemay be used including for example Hylon V (National Starch FoodInnovation) which has an amylose percentage of 56% or Hylon VII whichhas an amylose percentage of 70%. The remaining percentage isamylopectin. The polysaccharides pectin, amylose and sodium alginate areparticularly preferred for achieving colon delivery of the activeingredient.

It has been found that water-soluble polysaccharide, suitably pectin,can act as a former of pores in the coating otherwise provided byethylcellulose (preferably Surelease). By “pores” is not meantshaft-like holes from the surface to the core of the composition, ratherareas of weakness or absence of coating occurring stochastically on andwithin the coating of the invention.

Pore formers have been described before in connection with Surelease(see e.g. US 2005/0220878).

According to a particular embodiment of the invention the delayedrelease coating comprises ethylcellulose, e.g. Surelease™, and awater-soluble polysaccharide (WSP) wherein the proportion ofethylcellulose (in particular Surelease™) to WSP is ideally in the range90:10 to 99:1, preferably, 95:5 to 99:1, more preferably 97:3 to 99:1,for example about 98:2 based upon the dry weight of the coating.Suitably in this embodiment the weight gain of the composition due toapplication of the coating comprising ethylcellulose, e.g. Surelease™,and the WSP is in the range of from 1 to 30% (for example from: 3% to25%; 5% to 15%; 8% to 14%; 10% to 12%; 12% to 18%; or 16% to 18%,suitably the weight gain is about 11%, about 11.5%, or about 17%). It isparticularly preferred that the WSP in this embodiment is pectin.Particularly favoured weight gains using coatings comprisingethylcellulose, e.g. Surelease™, are those in the range 5-12% or in therange 8-12%.

Accordingly in an embodiment the second coating comprises ethylcellulose and a water soluble polysaccharide (particularly pectin)wherein the water-soluble polysaccharide (WSP) is present in an amountof 0.1% to about 10% by weight, based on the dry weight of the secondcoating. Suitably the WSP is present in an amount of from about 0.5% toabout 10%, for example about 0.5% to about 5%, about 1% to about 3%,suitably about 2% based on the total dry weight of the second coating.In this embodiment the WSP is preferably pectin. In this embodiment thesecond composition suitably further comprises a plasticizer. Suitableplasticizers include these described above in relation to Surelease™.Suitably the weight gain of the composition due to application of thesecond coating in this embodiment is in the range of from 1 to 30% (forexample from: 3% to 25%; 5% to 15%; 8% to 14%; 10% to 12%; 12% to 18%;or 16% to 18%, suitably the weight gain is about 11%, about 11.5%, orabout 17%).

In an embodiment the delayed release polymer is not a water-solublecellulose ether. Where the second coating comprises or is a delayedrelease polymer the delayed release polymer may not be the same as thewater-soluble cellulose ether of the first coating. Accordingly thesecond coating may not be the same as the first coating.

Continuous Phase Polymer Matrix (Aqueous Phase)

This section of the specification refers to a polymer matrix andcontinuous phase both of which concern the hydrogel forming polymermatrix. Therefore, reference to a polymer matrix or continuous phase canbe equated to the hydrogel forming polymer matrix. Furthermore, thissection of the specification relating to the polymer matrix recitesamounts of constituents in terms of percent by weight of thecomposition. In the context of this section of the specification, whatis meant is percent by weight of the dry weight of the composition orcore excluding coating(s).

The composition or the core may comprise a matrix or continuous phaseand also a disperse phase or oil phase. Similarly the liquid compositionof the invention comprises an aqueous phase comprising a hydrogelforming polymer. Suitably the continuous phase or matrix phase of thecomposition or core is or comprises a hydrogel-forming polymer. Ahydrogel-forming polymer is a polymer capable of forming a hydrogel. Ahydrogel may be described as a solid or semi-solid material, whichexhibits no flow when at rest, comprising a network (matrix) ofhydrophilic polymer chains that span the volume of an aqueous liquidmedium. A hydrogel forming polymer matrix is a network of hydrogelforming polymer chains, thus a hydrogel forming polymer matrix is ahydrogel forming polymer that has been allowed or caused to form amatrix.

The composition or core may comprise a hydrogel-forming polymer matrixand the liquid composition may select a hydrogel-forming polymerselected from the group consisting of: gelatin; agar; agarose; pectin;carrageenan; chitosan; alginate; starch; xanthan gum; gum Arabic; guargum; locust bean gum; polyurethane; polyether polyurethane; cellulose;cellulose ester, cellulose acetate, cellulose triacetate; cross-bondedpolyvinyl alcohol; polymers and copolymers of acrylic acid, hydroxyalkylacrylates, hydroxyethyl acrylate, diethylene glycol monoacrylate,2-hydroxypropylacrylate, 3-hydroxypropyl acrylate; polymers andcopolymers of methacrylic acid, hydroxyethyl methacrylate,diethyleneglycol monomethacrylate, 2-hydroxypropyl methacrylate,3-hydroxypropyl methacrylate, dipropylene glycol monomethylacrylate;vinylpyrrolidone; acrylamide polymers and copolymers,N-methylacrylamide, N-propylacrylamide; methacrylamide polymers andcopolymers, N-isopropylmethacrylamide, N-2-hydroxyethylmethacrylamide;and vinyl pyrrolidone; and combinations thereof. In specific embodimentsbinary or tertiary etc combinations of any of the above substances areforeseen.

In a further embodiment the hydrogel-forming polymer or the hydrogelforming polymer matrix is selected from the group consisting of gelatin,agar, a polyethylene glycol, starch, casein, chitosan, soya beanprotein, safflower protein, alginates, gellan gum, carrageenan, xanthangum, phthalated gelatin, succinated gelatin, cellulosephthalate-acetate,oleoresin, polyvinylacetate, polymerisates of acrylic or methacrylicesters and polyvinylacetate-phthalate and any derivative of any of theforegoing; or a mixture of one or more such hydrogel-forming polymers.

The hydrogel-forming polymer or the hydrogel forming polymer matrix mayalso be referred to as a hydrocolloid i.e. a colloid system wherein thecolloid particles are disperse in water and the quantity of wateravailable allows for the formation of a gel. In embodiments it ispreferred to use reversible hydrocolloids preferably thermo-reversiblehydrocolloids (e.g. agar, agarose, gelatin etc) as opposed toirreversible (single-state) hydrocolloids. Thermo-reversiblehydrocolloids can exist in a gel and sol state, and alternate betweenstates with the addition or elimination of heat. Gelatin, agar andagarose are thermo-reversible, rehydratable colloids and areparticularly preferred. Gelatin derivatives such as, for example,succinated or phthalated gelatins are also contemplated.Thermoreversible hydrocolloids which may be used according to theinvention, whether individually or in combination, include those derivedfrom natural sources such as, for example, carrageenan (extracted fromseaweed), gelatin (extracted from bovine, porcine, fish or vegetalsources), agar (from seaweed), agarose (a polysaccharide obtained fromagar) and pectin (extracted from citrus peel, apple and other fruits). Anon-animal based hydrocolloid may be preferred for certain applicationse.g. administration to vegetarians or to individuals not wishing toingest animal products for religious or health reasons. In relation tothe use of carrageenan, reference is made to US patent application2006/0029660 A1 (Fonkwe et al), the entirety of which is incorporatedherein by reference. The hydrogel-forming polymer may comprise or be acombination of gelatin with one or more other thermoreversiblehydrocolloids, e.g. with one or more other of the thermoreversiblehydrocolloids just listed. The hydrogel-forming polymer may comprise orbe a combination of gelatin with agar; optionally, at least one furtherthermoreversible hydrocolloid may be included in the combination, forexample one just listed.

Thermo-reversible colloids present a benefit over other hydrogel-formingpolymers. Gelation or hardening of thermo-reversible colloids occurs bycooling the colloid, e.g. in a liquid cooling bath or by air flow.Gelation of other hydrogel-forming polymers, which is chemically driven,can lead to leakage of the composition contents into the gelation mediumas the hardening process can take time to occur. Leakage of the contentof the composition may lead to an inaccurate quantity of the activeingredient within the composition. Thermo-reversible colloids are alsoknown as thermo-reversible gels, and it is therefore preferred that thehydrogel former be a thermo-reversible gelling agent.

Another term which may be applied to hydrogel formers which areadvantageous is “thermotropic”: a thermotropic gelling agent (which thereader will infer is preferred as a hydrogel former used in theinvention) is one caused to gel by a change in temperature and suchgelling agents are able to gel more rapidly than those whose gelling ischemically induced, e.g. ionotropic gelling agents whose gelling isinduced by ions, for example chitosan. In embodiments of the invention,therefore, the hydrogel former is a thermotropic gel-forming polymer ora combination of such polymers.

The manufacture of the composition to prepare a core may require thatthe hydrogel-forming polymer be present as a solution, which ispreferably an aqueous solution. The hydrogel-forming polymer representsbetween 5% and 50%, preferably between 10% and 30%, still morepreferably between 15% and 20% by weight of the aqueous phase duringmanufacture as described herein. In addition the hydrogel-formingpolymer may comprise 8 to 35%, (for example 15-25%, preferably 17-18%)hydro-gel forming polymer; 65%-85% (preferably 77-82%) of water plus,optionally, from 1-5% (preferably 1.5 to 3%) sorbitol. When presentsurfactant (e.g. anionic surfactant) in the aqueous phase pre-mix may bepresent in an amount of 0.1 to 5% (preferably 0.5 to 4%) wherein allparts are by weight of the aqueous phase.

In the aspect of the invention where the liquid composition is providedthe hydrogel forming polymer may be present as an aqueous solution inthe aqueous phase. The amounts of hydrogel forming polymer recited inthe immediately preceding paragraph are equally relevant to embodimentsof the liquid composition.

In embodiments the composition comprises at least 25%, suitably at least40% by weight based upon the dry weight of the composition of thehydrogel-forming polymer matrix. For example the hydrogel-formingpolymer matrix is present from 25 to 70%, for example 40 to 70% suitably45 to 60% of the composition, wherein the % is by weight based upon thedry weight of the composition.

In embodiments the hydrogel-forming polymer is a pharmaceuticallyacceptable polymer.

In certain embodiments the hydrogel-forming polymer is gelatin. Incertain embodiments the hydrogel-forming polymer matrix is gelatin. Incertain embodiments the hydrogel-forming polymer comprises gelatin. Incertain embodiments the gelatin comprises at least 30%, for example 30to 70% or 40 to 70% suitably 40 to 60% of the composition, wherein the %is by weight based upon the dry weight of the composition.

The hydrogel-forming polymer may optionally comprise a plasticiser forexample sorbitol or glycerine, or a combination thereof. In particularone or more plasticisers may be combined with gelatin.

In embodiments in which the hydrogel-forming polymer comprises or isgelatin, reference is hereby made to “Bloom strength”, a measure of thestrength of a gel or gelatin developed in 1925 by O. T. Bloom. The testdetermines the weight (in grams) needed by a probe (normally with adiameter of 0.5 inch) to deflect the surface of the gel 4 mm withoutbreaking it. The result is expressed in Bloom (grades) and usuallyranges between 30 and 300 Bloom. To perform the Bloom test on gelatin, a6.67% gelatin solution is kept for 17-18 hours at 10° C. prior to beingtested.

When the hydrogel-forming polymer comprises or is gelatin the bloomstrength of the gelatin may be in the range of 125 Bloom to 300 Bloom,200 Bloom to 300 Bloom and preferably 225 Bloom to 300 Bloom. It shouldbe appreciated that higher bloom strength gelatin can be replaced bylower bloom strength gelatin at higher concentrations.

According to the invention, in embodiments in which the hydrogel formingpolymer or hydrogel-forming polymer matrix comprises or is gelatin, thegelatin may be sourced by a variety of means. For example, it can beobtained by the partial hydrolysis of collagenous material, such as theskin, white connective tissues, or bones of animals. Type A gelatin isderived mainly from porcine skins by acid processing, and exhibits anisoelectric point between pH 7 and pH 9, while Type B gelatin is derivedfrom alkaline processing of bones and animal (bovine) skins and exhibitsan isoelectric point between pH 4.7 and pH 5.2. Type A gelatin issomewhat preferred. Gelatin for use in the invention may also be derivedfrom the skin of cold water fish. Blends of Type A and Type B gelatinscan be used in the invention to obtain a gelatin with the requisiteviscosity and bloom strength characteristics for bead manufacture.

Lower temperature gelatin (or gelatin derivatives or mixtures ofgelatins with melting point reducers) or other polymer matrices able tobe solidified at lower temperatures (e.g. sodium alginate) may also beused. It is therefore believed that polymer which comprises or is lowtemperature gelatin is a preferred matrix polymer.

According to the invention, in embodiments in which the hydrogel formingpolymer or hydrogel forming polymer matrix comprises or is gelatin, thestarting gelatin material is preferably modified before manufacture toproduce “soft gelatin” by the addition of a plasticizer or softener tothe gelatin to adjust the hardness of the composition of the invention.The addition of plasticizer achieves enhanced softness and flexibilityas may be desirable to optimise dissolution and/or further processingsuch as, for example, coating. Useful plasticizers of the presentinvention for combination with gelatin or another hydrogel-formingpolymer include glycerine (1,2,3-propanetriol), D-sorbitol (D-glucitol),sorbitol BP (a non-crystallizing sorbitol solution) or an aqueoussolution of D-sorbitol, sorbitans (e.g. Andidriborb 85/70), mannitol,maltitol, gum arabic, triethyl citrate, tri-n-butyl citrate,dibutylsebacate. Other or similar low molecular weight polyols are alsocontemplated for example ethylene glycol and propylene glycol.Polyethylene glycol and polypropylene glycol may also be used althoughthese are less preferred. Glycerine and D-sorbitol may be obtained fromthe Sigma Chemical Company, St. Louis, Mo. USA or Roquette, France. Someactive agents and excipients included for other functions may act asplasticisers.

Softeners or plasticisers, if utilized, can be ideally incorporated in aproportion rising to 30%, preferably up to 20% and more preferably up to10% by dry weight of the composition of the invention, even morepreferably between 3 and 8%, and most preferably between 4% and 6%.

Although not essential, the hydrogel-forming polymer matrix may alsooptionally contain a disintegrant where it is particularly desired toenhance the rate of disintegration of the composition of the invention.Examples of disintegrants which may be included are alginic acid,croscarmellose sodium, crospovidone, low-substituted hydroxypropylcellulose and sodium starch glycolate.

A crystallisation inhibitor (e.g. approximately 1% by dry weight of thecomposition) may also be included in the composition of the invention.An example is hydroxy propyl/methyl cellulose (HPC or HPMC, hypromelloseetc) which may play other roles such as, for example, emulsifier.

In another embodiment, the hydrogel-forming polymer matrix is chitosanwhich can exist in the form of biogels with or without additives asdescribed e.g. in U.S. Pat. No. 4,659,700 (Johnson & Johnson); by KumarMajeti N.V. Ravi in Reactive and Functional Polymers, 46, 1, 2000; andby Paul et al. in ST. P. Pharma Science, 10, 5, 2000 the entirety of all3 of which is incorporated herein by reference. Chitosan derivativese.g. thiolated entities are also contemplated.

The hydrogel-forming polymer matrix may be a non-hydrocolloid gum.Examples are the cross-linked salts of alginic acid. For example,aqueous solutions of sodium alginate gums extracted from the walls ofbrown algae have the well known property of gelling when exposed to di-and trivalent cations. A typical divalent cation is calcium, often inthe form of aqueous calcium chloride solution. It is preferred in thisembodiment that the cross-linking or gelling have arisen throughreaction with such a multivalent cation, particularly calcium.

The hydrogel-forming polymer matrix may have a low water content,therefore the composition may have a low water content. As describedbelow, during manufacture of a core the disperse phase or oil phase,optionally comprising cyclosporin, is mixed with an aqueous solution ofthe hydrogel-forming polymer and the composition is gelled, for exampleto provide a composition or a core which are minibeads. Suitably thecomposition or cores are dried following formation to reduce the watercontent present therein.

In certain embodiments the composition does not comprise compoundscontaining a disulphide bond. In embodiments the hydrogel-formingpolymer does not comprise compounds containing a disulphide bond.

The hydrogel-forming polymer matrix forming the continuous phase of thecore (aqueous phase) may further comprise a surfactant. Surfactantswhich may be used in the composition are described in the section“surfactants” below.

Surfactant which may be present in the continuous phase, aqueous phaseor the hydrogel forming polymer matrix of the composition or coreinclude, for example a surfactant selected from the group consisting of:cationic; amphoteric (zwitterionic); anionic surfactants, for exampleperfluoro-octanoate (PFOA or PFO), perfluoro-octanesulfonate (PFOS),sodium dodecyl sulfate (SDS), ammonium lauryl sulfate, and other alkylsulfate salts, sodium laureth sulfate, also known as sodium lauryl ethersulfate (SLES) and alkyl benzene sulphonate; and non-ionic surfactantsfor example perfluorocarbons, polyoxyethyleneglycol dodecyl ether (e.g.Brij such as, for example, Brij 35), Myrj (e.g. Myrj 49, 52 or 59),fatty alcohol ethoxylates, alkylphenol ethoxylate, fatty acidethoxylates, fatty amide ethoxylates, alkyl glucosides, Tween 20 or 80(also known as Polysorbate) (Brij, Myrj and Tween products are availablecommercially from Croda), poloxamers which are nonionic triblockcopolymers composed of a central hydrophobic chain of polyoxypropylene(poly(propylene oxide)) flanked by two hydrophilic chains ofpolyoxyethylene (poly(ethylene oxide)), or a combination of theforegoing. In particular, the surfactant may be selected from, orcomprise, anionic surfactants and combinations thereof, the anionicsurfactants optionally being those mentioned in this paragraph. Aparticular class of surfactant comprises sulfate salts. A preferredanionic surfactant in the aqueous phase is SDS. Mixtures of anionicsurfactants may be used. Mixtures of further surfactants are alsocontemplated, e.g. mixtures comprising perfluorocarbons.

In embodiments of the invention, the core comprises a hydrophilicsurfactant which, without being bound by theory, is believed at leastpartially to partition the aqueous phase (polymer matrix).

Such surfactants intended for such inclusion in the aqueous phase of thecore are preferably readily diffusing or diffusible surfactants tofacilitate manufacturing and processing of the composition of theinvention.

The surfactant may have an HLB of at least 10 and optionally of at least15, e.g. at least 20, or at least 30 and optionally of 38-42, e.g. 40.Such surfactants can be of any particular type (ionic, non-ionic,zwitterionic) and may comprise as a proportion of dry weight of thecomposition from 0.1% to 6%, e.g. 0.1% to 5%. 0.1% to 4% or 0.1% to 3%,more preferably in a proportion of at least 1% and in particular between1.0 and 4.5 or 5%, ideally within or just outside the 2-4% range, forexample from 2 to 3% or approximately 2% or approximately 4%.

Unless otherwise stated or required, all percentages and ratios are byweight.

In one embodiment the anionic surfactant which may be present in thecontinuous phase, aqueous phase or the hydrogel forming polymer matrixof the composition or core may be an anionic surfactant selected fromalkyl sulphates, carboxylates or phospholipids, or combinations thereof.

The physical form of the surfactant at the point of introduction intothe aqueous phase during preparation of the composition or core plays arole in the ease of manufacture of the composition or core. As such,although liquid surfactants can be employed, it is preferred to utilizea surfactant which is in solid form (e.g. crystalline, granules orpowder) at room temperature, particularly when the aqueous phasecomprises gelatin.

Disperse Phase

The polymer matrix or the continuous phase of the composition, or inembodiments where a core is present, the core described above (forexample the hydrogel-forming polymer) may comprise a disperse phase.Similarly the aqueous phase of the liquid composition comprises adisperse phase which is or comprises the oil phase. Suitably thedisperse phase, where present, may comprise the cyclosporin. In suchembodiments the cyclosporin is preferably soluble in the disperse phase,i.e. the disperse phase comprises a vehicle in which the active isdissolved. Embodiments wherein the cyclosporin is solubilised in thedisperse phase are preferred, because such compositions release thecyclosporin in a solubilised form, which may enhance the therapeuticeffect of the drug at the site of release, for example by enhancingabsorption into the colonic mucosa.

In embodiments the cyclosporin is or is comprised in the disperse phase.The disperse phase is or comprises the oil phase. Preferably, thedisperse phase is the oil phase.

The disperse phase may comprise a water immiscible phase (also referredto herein as an oil phase). The water immiscible phase may be solid,semi-solid or liquid at ambient temperature (e.g. 25° C.), and thereforethe oil phase may for example be waxy at ambient temperature. The oilphase may be or may comprise a liquid lipid and optionally a solventmiscible therewith. The cyclosporin may be present in the oil phase.Suitably the cyclosporin is soluble in the oil phase.

The disperse phase may comprise a combination of oils, for exampleliquid lipids. The liquid lipid may be a short-, medium- or long-chaintriglyceride formulation, or a combination thereof. A medium chaintriglyceride(s) (MCT) comprises one or more triglycerides of at leastone fatty acid selected from C₆, C₇, C₈, C₉, C₁₀, C₁₁ and C₁₂ fattyacids. It will be understood that commercially available triglyceride,in particular MCT, formulations useful in the invention are mixturesderived from natural products and usually or always contain minoramounts of compounds which are not MCTs; the term “medium chaintriglyceride formulation” is therefore to be interpreted to include suchformulations. A short chain triglyceride(s) comprises one or moretriglycerides of at least one short chain fatty acid selected from C₂-C₅fatty acids. A long chain triglyceride(s) comprises one or moretriglycerides of at least one long chain fatty acid having at least 13carbon atoms.

The liquid lipid may comprise or be triglycerides and/or diglycerides.Such glycerides may be selected from medium chain triglycerides or shortchain triglycerides or a combination thereof.

The liquid lipid may be a caprylic/capric triglyceride, i.e. acaprylic/capric triglyceride formulation (which it will be understoodmay contain minor amounts of compounds which are not caprylic/caprictriglycerides).

The disperse phase may optionally comprise a solvent. Accordingly theoil phase may comprise a solvent. Said solvent which is optionallyincluded in an oil phase may be miscible with both the liquid lipid andwith water. Examples of suitable solvents are 2-(2-ethoxyethoxy)ethanolavailable commercially under trade names Carbitol™, Carbitol cellosolve,Transcutol™, Dioxitol™, Poly-solv DE™, and Dowanal DE™; or the purerTranscutol™ HP (99.9). Transcutol P or HP, which are availablecommercially from Gattefosse, are preferred. Another possible co-solventis poly(ethylene glycol). PEGs of molecular weight 190-210 (e.g. PEG200) or 380-420 (e.g. PEG 400) are preferred in this embodiment.Suitable PEGs can be obtained commercially under the name “Carbowax”manufactured by Union Carbide Corporation although many alternativemanufacturers or suppliers are possible.

The disperse phase may represent from 10-85% by dry weight of the core.

As discussed above the disperse phase may be an oil phase comprising anypharmaceutically suitable oil, e.g. a liquid lipid. The oil phase may bepresent as oil drops. In terms of dry weight of the core, the oil phasemay comprise a proportion from 10% to 85%, e.g. 15% to 50%, for example20% to 30% or from 35% to 45%. The term “oil” means any substance thatis wholly or partially liquid at ambient temperature or close-to-ambienttemperature e.g. between 10° C. and 40° C. or between 15° C. and 35° C.,and which is hydrophobic but soluble in at least one organic solvent.Oils include vegetable oils (e.g. neem oil) and petrochemical oils.

The oil may be present in the composition in an amount of from about 2%to about 25%, from about 3% to about 20%, from about 3% to about 10% orfrom about 5% to about 10% by weight based upon the dry weight of thecore.

Oils which may be included in the oil phase include poly-unsaturatedfatty acids such as, for example, omega-3 oils for exampleeicosapentanoic acid (EPA), docosohexaenoic acid (DHA), alpha-linoleicacid (ALA), conjugated linoleic acid (CLA). Preferably ultrapure EPA,DHA or ALA or CLA are used e.g. purity up to or above 98%. Omega oilsmay be sourced e.g. from any appropriate plant e.g. sacha inchi. Suchoils may be used singly e.g. EPA or DHA or ALA or CLA or in anycombination. Combinations of such components including binary, tertiaryetc combinations in any ratio are also contemplated e.g. a binarymixture of EPA and DHA in a ratio of 1:5 available commercially underthe trade name Epax 6000. The oil part of the oil phase may comprise orbe an oil mentioned in this paragraph.

Oils which may be included in the oil phase are particularly naturaltriglyceride-based oils which include olive oil, sesame oil, coconutoil, palm kernel oil, neem oil. The oil may be or may comprise saturatedcoconut and palm kernel oil-derived caprylic and capric fatty acids andglycerin e.g. as supplied under the trade name Miglyol™ a range of whichare available and from which one or more components of the oil phase ofthe invention may be selected including Miglyol™ 810, 812(caprylic/capric triglyceride); Miglyol™ 818: (caprylic/capric/linoleictriglyceride); Miglyol™ 829: (caprylic/capric/succinic triglyceride;Miglyol™ 840: (propylene glycol dicaprylate/dicaprate). Note thatMiglyol™ 810/812 are MCT formulations which differ only in C₈/C₁₀-ratioand because of its low C₁₀-content, the viscosity and cloud point ofMiglyol™ 810 are lower. The Miglyol™ range is available commerciallyfrom Sasol Industries. As noted above, oils which may be included in theoil phase need not necessarily be liquid or fully liquid at roomtemperature. Waxy-type oils are also possible: these are liquid atmanufacturing temperatures but solid or semi-solid at normal ambienttemperatures. The oil part of the oil phase may comprise or be an oilmentioned in this paragraph.

Alternative or additional oils which may be included in the oil phaseaccording to the invention are other medium chain triglycerideformulations such as for example Labrafac™ Lipophile manufactured byGattefosse in particular product number WL1349. Miglyol™ 810, 812 arealso medium chain triglyceride formulations.

Accordingly the oil phase may be or comprise medium chain mono-di- ortri-glycerides.

The medium chain glyceride(s) (eg mono- di- or tri-glyceride(s))mentioned herein are those which comprise one or more triglycerides ofat least one fatty acid selected from fatty acids having 6, 7, 8, 9, 10,11 or 12 carbon atoms, e.g. C8-C10 fatty acids.

The oil phase may further comprise the surfactant as described above andelsewhere herein. The presence of the surfactant in the oil phase mayalso provide a stabilising effect on the liquid composition when the oilphase is dispersed in the aqueous phase. In addition the presence of thesurfactant in the oil phase may inhibit crystallisation of cyclosporinfrom a cyclosporin solution in the oil phase. The surfactant may alsoprovide enhanced emulsification when the disperse phase is mixed withthe aqueous phase during preparation of the liquid composition,composition or core (i.e act as an emulsifier).

The liquid lipid or oil of the oil phase or disperse phase is suitablynot a surfactant. However, certain oils, particularly those derived fromnatural sources will comprise components which may have surface activeproperties. For example many triglyceride oils also comprise mono anddiglyceride components and may therefore exhibit some surfactant likeproperties. Accordingly the oil suitably has an HLB value of 0-10,however suitably the oil has an HLB which is close to 0 for example anHLB of 0 to 3, optionally about 0, about 1 or about 2.

Surfactant in the oil phase may for example be or comprise a mediumchain or long chain fatty acid mono- or di-glyceride or a combinationthereof, wherein the surfactant does not comprise or is not apolyethyleneglycol ether or ester. Optionally the surfactant is a mediumchain or long chain fatty acid mono-glyceride, di-glyceride or acombination thereof, optionally wherein the surfactant does not compriseor is not a polyethyleneglycol ether or ester. Two particularsurfactants contemplated by the invention are glyceryl caprylate/caprateand glyceryl monooleate/dioleate. Commercial preparations may also beused as a surfactant e.g. those commercial preparations which containminor components. Preferred examples are Capmul GMO-50 (glycerylmonooleate/dioleate) and Capmul MCM (glyceryl caprylate/caprate).

Within embodiments, the HLB of the oil may be in the range 0-10(optionally 1-8, e.g. 1-6 and sometimes 1-5).

In another embodiment the oil phase comprises an oil with an HLB in therange 0-10 (preferably 1-5) and the has an HLB of up to 10 andoptionally up to 7, 1-8, 1-7, 1-5, 2-5, 1-4, 1-3, 1-2, 2-4, 3-4, 5-8,6-8 and 6-7.

In another embodiment the oil phase comprises an oil and the surfactantwherein the oil and the surfactant both have an HLB in the range 0-10.For example the oil has an HLB of 1-5, for example 1 to 4 or 1-2 and thesurfactant has an HLB 2-8, for example 3-7, 2-6, or 3-4).

Suitable oils which may comprise or be the oil phase or disperse phasewith a low HLB (HLB less than 10) include medium chain triglycerides,caprylocaproyl macrogolglycerides and caprylic/capric triglyceride. Interms of commercial products, particularly preferred oils in the lowerHLB range are Labrafac™ Lipophile (e.g. 1349 WL), Captex 355 and Miglyol810.

It is to be understood that the oil phase or disperse phase in theembodiments above may further comprise one or more solvents, for example2-(2-ethoxyethoxy)ethanol or low molecular weight PEG as mentionedabove. The solvent may be present in the composition in an amount ofform about 1% to 30%, for about 5% to about 30%, for about 10% to about25%, or from about 12% to about 22% by weight based upon the dry weightof the uncoated composition or upon the dry weight of the core.

A particular oil phase comprises an oil (low HLB), the surfactant and aco-solvent. For example the following three commercial products:Transcutol P (as co-solvent), Myglyol 810 (as oil) and Capmul GMO-50(surfactant). An oil phase may therefore comprise or consist of acombination of the following: 2-ethoxyethanol, an MCT and particularly acaprylic/capric triglyceride formulation, and glycerylmonooleate/dioleate. The oil phase may further comprise the cyclosporin.

Preferably, cyclosporin is soluble in the oil phase. As discussed belowin relation to preparation of the composition, the cyclosporin issuitably dissolved in the oil phase and the oil phase is mixed with anaqueous phase comprising the hydrogel-forming polymer.

The disperse phase (oil phase) may be or comprise a glycerideformulation, optionally wherein the disperse phase is or comprises afatty acid monoglyceride, diglyceride or triglyceride or a combinationthereof, or the disperse phase is or comprises a caprylic/caprictriglyceride formulation.

The disperse phase of the colloidal core may comprise self-assemblystructures, for example micelles, vesicles, liposomes or nanoparticles,or at least the structures which result from drying aqueous colloids ofsuch types (have the characteristics of structures which result fromdrying aqueous colloids of such types). The invention in particularincludes formulations in which the disperse phase is micellar, i.e.formed of micelles and/or promicelles. The term “promicelle” refers to apart of a formulation which will form a micelle upon contact with water,e.g. gastrointestinal contents.

The following discussion for convenience refers to micelles but isapplicable in general to other self-assembly structures. Amicelle-forming surfactant is present as micelles dispersed within thehydrogel-forming polymer in a “wet” (not yet dried) composition made asan intermediate in the manufacturing process described herein. It isbelieved also to be present as micelles in the dried composition butobservability of micelles or micelle-like structures in the driedcomposition is not a requirement of the invention. It is mentioned atthis point that the presence of a surfactant in micelle form does notrequire that the entire surfactant content of a composition is inmicelle form as it is considered more probable that a portion of thesurfactant will be outside the micelles. Thus in the “wet” composition,whether the hydrogel-forming polymer is in the gel state or the sol(liquid) state may comprise the micelle-forming surfactant at aconcentration above the critical micelle concentration.

The diameter of the dispersed micelles may be between 0.5 nm and 200 nm,1 nm and 50 nm, or 5 nm and 25 nm. The size of the micelles may bedetermined by dynamic light scattering or diffusion NMR techniques knownwithin the art. Although the size of the micelles is given as a diameterthis does not imply that the micelles must be purely spherical speciesonly that they may possess some approximately circular dimension.

The surfactant may be a non-ionic surfactant. The surfactant may be apolyoxyethylated surfactant. The surfactant has a hydrophilic head whichmay be a hydrophilic chain, for example a polyoxyethylene chain or apolyhydroxylated chain.

The surfactant of course has a hydrophobic part and in particular ahydrophobic chain. The hydrophobic chain may be a hydrocarbon chain, forexample having at least 6 carbon atoms and optionally at least 10 carbonatoms, and particularly of at least 12 carbon atoms; some hydrocarbonchains have no more than 22 carbon atoms, for example C₁₀-C₂₀, C₁₂-C₂₀or C₁₅-C₂₀ hydrocarbon chains. It may be an alkyl chain, e.g. having anumber of carbon atoms just mentioned. It may be an alkenyl chaincomprising one or more carbon-carbon double bonds, e.g. having a numberof carbon atoms just mentioned. The surfactant may comprise ahydrocarbon chain, e.g. alkyl chain or alkenyl chain that is substitutedprovided that it maintains a hydrophobic characteristic. There may forexample be one or two substituents, for example a single substituent,e.g. selected from halogen (e.g. F or Cl), hydroxy, thiol oxo, nitro,cyano; hydroxy or thiol substituents may be esterified by for example afatty acid. One class of surfactants comprise a hydrocarbonmonosubstituted by hydroxy; optionally, at least a portion of thehydroxy groups of an aliquot of surfactant, e.g. of the surfactant in abead, may be esterified by a fatty acid or mono-hydroxy fatty acid asdisclosed herein or etherified by a fatty alcohol for example having atleast 6 carbon atoms and optionally at least 10 carbon atoms, andparticularly of at least 12 carbon atoms; some hydrocarbon chains haveno more than 22 carbon atoms, for example C₁₀-C₂₀, C₁₂-C₂₀ or C₁₅-C₂₀fatty alcohols.

The hydrophobic chain may be part of an esterified fatty acid R¹—COOH orof an etherified or esterified fatty ether R¹—COH where R¹ is thehydrophobic chain, e.g. as mentioned in the preceding paragraph. Theester-forming or, as the case may be, ether-forming group will typicallycomprise a hydrophilic chain.

As mentioned, the surfactant may have a hydrophilic chain and may be anon-ionic surfactant, and may satisfy both requirements. The hydrophilicchain may be a poly(ethyleneglycol), also known as poly(oxyethylene) ormacrogol. The hydrophilic chain may be of the formula—(O—CH₂—CH₂)_(n)—OR where n is 5 or 6 to 50 and R is H or alkyl, e.g.ethyl or methyl. The invention includes implementations in which n isfrom 6 to 40, e.g. from 6 to 35. In some embodiments, n is from 6 to 25and optionally is from 8 to 25 or from 8 to 15. In other embodiments, nis from 8 to 50 or from 8 to 40, e.g. is from 10 to 50, 10 to 40 or 10to 35. In a particular embodiment, n is 15. For all hydrophilic chainsof the formula —(O—CH₂—CH₂)_(n)—OR, in one class of embodiments R is H.

The hydrophilic chain may be a polyhydroxylated chain (for example aC₅-C₂₀ e.g. C₅-C₁₀ chain), e.g. having a hydroxy group on the carbonatoms of the chain, for example a glucamide.

The micelle-forming surfactant may comprise a combination of ahydrophobic chain as described above and a hydrophilic chain asdescribed above. It may therefore be, or comprise, a macrogol ester of afatty acid as described herein or a macrogol ether of a fatty alcohol asdescribed herein.

Micelle-forming surfactants comprising a hydrophobic chain and ahydrophilic chain can be selected from the group consisting of: macrogolesters; macrogol ethers; diblock copolymers; triblock copolymers; andamphiphilic polymers. In certain embodiments of the invention anycombinations of the group are included within the invention.

Examples of macrogol esters which are suitable for use in the presentinvention are macrogol esters of fatty acids having at least 6 carbonatoms and optionally at least 10 carbon atoms, and particularly of atleast 12 carbon atoms; some fatty acids have no more than 22 carbonatoms, for example C₁₀-C₂₀, C₁₂-C₂₀ or C₁₅-C₂₀ fatty acids. The fattyacids may be saturated or unsaturated but are in particular saturated.To be mentioned are macrogol 25 cetostearyl ether (Cremophor® A25);macrogol 6 cetostearyl ether (Cremophor® A6); macrogol glycerolricinoleate 35 (Cremophor® EL); macrogol-glycerol hydroxystearate 40(Cremophor® RH 40); macrogol-15-hydroxystearate (Solutol® HS 15).Examples of macrogol ethers which are suitable for use in the presentinvention are macrogol ethers of fatty alcohols having at least 6 carbonatoms and optionally at least 10 carbon atoms, and particularly of atleast 12 carbon atoms; some fatty alcohols have no more than 22 carbonatoms, for example C₁₀-C₂₀, C₁₂-C₂₀ or C₁₅-C₂₀ fatty alcohols. The fattyalcohols may be saturate or unsaturated but are in one embodimentsaturated.

Examples of amphiphilic polymers which are suitable for use in thepresent invention are: alkyl glucamides; fatty alcohol poly(ethoxyl)atesalso known as polyethoxylated alkyl ethers; poly(ethoxyl)ated fatty acidesters (Myrj or Solutol); fatty amide polyethoxylate; fatty amineethoxylate; alkylphenol ethoxylate; polyethoxylated sorbitan esters(polysorbates); polyethoxylated glycerides; or poly-glycerol esters.

Examples of copolymers, which are suitable for use in the presentinvention are: pluronics(poloxamers);polyvinylpyrollidone-polyvinylacetate (Plasdone S630); aminoalkylmethacrylate copolymer (Eudragit EPO); methacrylic acid-methylmethacrylate copolymer (Eudragit S100, L100); polycaprolactone-PEG;polycaprolactone-methoxy-PEG; poly(aspartic acid)-PEG;poly(benzyl-L-glutamate)-PEG; poly(D,L-lactide)methoxy-PEG;poly(benzyl-L-aspartate-PEG; or poly(L-lysine)-PEG

In a preferred embodiment the micelle-forming surfactant cis a macrogolester, more preferably a macrogol ester that conforms to the EuropeanPharmacopoeia monograph number 2052 macrogol-15-hydroxystearate, such asKolliphor® HS 15 marketed by BASF.

Kolliphor® HS 15 consists of polyglycol mono- and di-esters of12-hydroxystearic acid and about 30% of free polyethylene glycol. Themain components of the ester part have the following chemicalstructures:

where x and y are integers and a small part of the 12-hydroxy group canbe etherified with polyethylene glycol.

Suitable surfactants comprise those which during manufacture combinewith the aqueous phase (including hydrogel-forming polymer) in an amountabove their CMC to form a clear liquid. Kolliphor® HS 15 is such asurfactant.

In certain embodiments the weight ratio of the micelle-formingsurfactant to the antigen is from 10:1 to 100:1, optionally from 50:1 to100:1. In some embodiments, the ratio is from 80:1 to 90:1. Inparticular embodiments, the ratio is from 50:1 to 60:1.

In particular embodiments, the compositions of the invention comprise acombination of micelle-forming compounds. Such a combination ofmicelle-forming compounds may consist of two or more surfactants asmentioned in the preceding section of this specification. Alternatively,a surfactant may be combined with one or more other compounds at leastpotentially able to form micelles with the surfactant, optionallyselected from cationic lipids and glycolipids, amongst others. As anadditional option, a composition may comprise a plurality of surfactantsas mentioned in the preceding section of this specification and one ormore other compounds at least potentially able to form micelles with thesurfactant, optionally selected from cationic lipids and glycolipids,amongst others.

The invention therefore includes compositions as described herein whichcomprise:

two or more micelle-forming surfactants, e.g. two or more surfactantshaving a hydrophobic chain and a hydrophilic chain;

a compound, e.g. a single compound or two or more compounds, selectedfrom cationic lipids and glycolipids;

two or more micelle-forming surfactants and a compound, e.g. a singlecompound or two or more compounds, selected from cationic lipids andglycolipids.

A disperse phase which is or comprises a surfactant may enhance theabsorption of an active ingredient, for example cyclosporin A, into thetissue of the GIT, for example by forming self-assembly structures, suchas micelles, which are associated with the active ingredient and thuspresent the drug to the mucosa tissue of the GI tract in a form whichenhances uptake/absorption into the tissue.

The oil phase may also include one or more volatile or non-volatilesolvents, which may be the same or different from the solvent orco-solvent previously mentioned. Such solvents may for example remain inthe formulation of the invention following processing e.g. initialdissolution of the components present in the core, and have noparticular function in the core formulation. Alternatively, suchsolvents if present may function to maintain the cyclosporin a dissolvedstate (in solution) within the oil phase or to facilitate dispersion,egress etc. In other embodiments, the solvent may have partly or fullyevaporated during processing and therefore be present in only minorquantities if at all. In a related embodiment, the solvent, particularlywhen a solvent which is both oil and water-soluble is used, may bepartly or completely present in the aqueous phase of the core. Anexample of such a solvent is ethanol. Another example is transcutolwhich is already mentioned as a co-solvent.

Accordingly, the composition may comprise a hydrogel-forming polymermatrix which forms a continuous phase and a disperse phase comprisingcyclosporin, a low HLB medium or long chain mono- or di-estersurfactant, a low HLB oil, and optionally a co-solvent. Optionally, themedium or long-chain mono- or di-ester surfactant is a medium- orlong-chain mono- or di-glyceride surfactant.

In a particular embodiment the composition or the core is in the form ofa solid colloid, the colloid comprising a continuous phase and adisperse phase, wherein the disperse phase is or comprises:

cyclosporin;

a medium chain mono-, di- and/or tri-glyceride, for example a mediumchain triglyceride, particularly caprylic/capric triglyceride;

a medium- or long-chain mono- or di-glyceride, particularly glycerylmonooleate/dioleate; and

a co-solvent (for example 2-(ethoxyethoxy)ethanol);

and wherein the continuous phase is or comprises:

a hydrogel-forming polymer matrix which is or comprises a hydrocolloidselected from carrageenan, gelatin, agar and pectin, or a combinationthereof optionally selected from gelatin and agar or a combinationthereof, more particularly the polymer of the a hydrogel-forming polymermatrix is or comprises gelatin;

a plasticiser, optionally a plasticiser selected from glycerin, a polyolfor example sorbitol, polyethylene glycol and triethyl citrate or amixture thereof, particularly sorbitol; and

an anionic surfactant, for example at least one surfactant selected fromfatty acid salts, alkyl sulphates and bile salts, particularly an alkylsulphate, for example sodium dodecyl sulphate.

In a further specific embodiment the disperse phase comprises:

cyclosporin in an amount of 60-180 mg/g;

caprylic/capric triglyceride in an amount of 40-80 mg/g;

2-(2-ethoxyethoxy)ethanol in an amount of 100-200 mg/g; and

glyceryl monooleate and/or glyceryl dioleate in an amount of 100-150mg/g, wherein weights are based upon the dry weight of the composition.

The oil phase or disperse phase may comprise:

cyclosporin in an amount of 120-360 mg/g;

caprylic/capric triglyceride in an amount of 80-160 mg/g;

2-(2-ethoxyethoxy) ethanol in an amount of 200-400 mg/g; and

glyceryl monooleate and/or glyceryl dioleate in an amount of 200-300mg/g,

wherein the weights are based on the weight of the wet composition.

The liquid composition may comprise an oil phase comprising:

cyclosporin in an amount of 20-60 mg/g;

caprylic/capric triglyceride in an amount of 13-27 mg/g;

2-(2-ethoxyethoxy) ethanol in an amount of 50-70 mg/g; and

glyceryl monooleate and/or glyceryl dioleate in an amount of 30-55 mg/g,

wherein weights are based upon the wet weight of the composition, i.e.the liquid composition, optionally wherein the oil phase to aqueousphase ratio may be 1:5.

In an embodiment the aqueous phase or continuous phase comprises ahydrogel-forming polymer matrix comprising gelatin in an amount of 300to 700 mg/g, and SDS in an amount of 15-50 mg/g, wherein weights arebased upon the dry weight of the composition.

In an embodiment the aqueous phase may comprise a hydrogel-formingpolymer matrix comprising gelatin in an amount of 120 to 280 mg/g andSDS in an amount of 6-20 mg/g wherein the weights are based upon theweight of the aqueous phase. The aqueous phase may comprise ahydrogel-forming polymer matrix comprising gelatin in an amount of 100to 230 mg/g and SDS in an amount of 5-16 mg/g, wherein the weights arebased on the weight of the composition, i.e. the liquid composition,optionally wherein the oil phase to aqueous phase ratio may be 1:5.

Suitably in the embodiment of the immediately preceding two paragraphsthe cyclosporin may be present in an amount of 90 to 140 mg/g, forexample of 60 to 150 mg/g, 80 to 120 mg/g or particularly 80 to 100mg/g. The anionic surfactants are as defined herein, for example ananionic surfactant selected from alkyl sulphates, carboxylates orphospholipids (particularly SDS).

The composition or the cores described above comprising hydrogel-formingpolymer matrix may be coated as described herein. A particular coatingfor these embodiments is a coating comprising:

a first coating (sub-coating) which is or comprises a water-solublecellulose ether, particularly hydroxypropylmethyl cellulose;

a second coating outside the first coating which is or comprises amodified release coating, particularly a pH independent modified releasecoating, more especially a coating comprising ethyl cellulose (e.g.Surelease) still more particularly a coating comprising ethyl celluloseand a water-soluble polysaccharide such as pectin (e.g. aSurelease-pectin coating as described herein); and wherein

the first coating is present in an amount corresponding to a weight gaindue to the first coating in a range selected from: (i) from 8% to 12%,for example about 10%; or (ii) from 4% to 6%, for example about 5% byweight based upon the weight of the formulation prior to applying thefirst coating; and wherein

the second coating is present in an amount corresponding to a weightgain of the formulation due to the second coating selected from (a) from10% to 12%, for example about 11% or about 11.5%; or (b) from 16% to18%, for example about 17% by weight based upon the weight of theformulation prior to applying the second coating.

Equally, the composition or the cores described above comprisinghydrogel-forming polymer matrix may be coated with a coating comprising:

a second coating which is or comprises a modified release coating,particularly a pH independent modified release coating, more especiallya coating comprising ethyl cellulose (e.g. Surelease) still moreparticularly a coating comprising ethyl cellulose and a water-solublepolysaccharide such as pectin (e.g. a Surelease-pectin coating asdescribed herein); and wherein

the second coating is present in an amount corresponding to a weightgain of the formulation due to the second coating selected from (a) from10% to 12%, for example about 11% or about 11.5%; or (b) from 16% to18%, for example about 17% by weight based upon the weight of theformulation prior to applying the second coating.

Surfactant

The composition comprises a surfactant, as described above. Thesurfactant may be present in the composition or the core, for example inthe hydrogel-forming polymer matrix, or in the disperse phase or both.The surfactant may also be present in one or more of the coatingscomprised in the composition or applied to the core.

The composition may comprise a further surfactant. Where the compositioncomprises a further surfactant this surfactant can be referred to as asecond surfactant and the surfactant present in the composition of theinvention can be referred to as a first surfactant. Accordingly, thefirst surfactant is or comprises the medium chain or long chain fattyacid mono- or di-glyceride or a combination thereof, which does notcomprise or is not a polyethyleneglycol ether or ester. The furthersurfactant may be present in the composition or the core, for example inthe hydrogel-forming polymer matrix, or in the disperse phase or both.The further surfactant may also be present in one or more of thecoatings comprised in the composition or applied to the core. Suitablefurther surfactants can be anionic, cationic, zwitterionic, ornon-ionic.

In the description and claims of this specification, the term“surfactant” is employed as a contraction for “surface active agent”.For the purposes of this description and claims, it is assumed thatthere are four major classifications of surfactants; therefore thefurther surfactant may be: anionic, cationic, non-ionic, and amphoteric(zwitterionic). The non-ionic surfactant remains whole, has no charge inaqueous solutions, and does not dissociate into positive and negativeions. Anionic surfactants are water-soluble, have a negative charge anddissociate into positive and negative ions when placed in water. Thenegative charge lowers the surface tension of water and acts as thesurface-active agent. Cationic surfactants have a positive charge, andalso dissociate into positive and negative ions when placed in water. Inthis case, the positive ions lower the surface tension of the water andact as the surfactant. The amphoteric (zwitterionic) surfactant assumesa positive charge in acidic solutions and performs as a cationicsurfactant, or it assumes a negative charge in an alkaline solution andacts as an anionic surfactant.

The further surfactant(s) may be selected from: anionic surfactants andcombinations thereof; from non-ionic surfactants and combinationsthereof; and from combination of an anionic surfactant (e.g. a singlesuch surfactant or a plurality thereof) and a non-ionic surfactant (e.g.a single such surfactant or a plurality thereof). Preferably the secondsurfactant is an anionic surfactant.

Accordingly, in an embodiment the liquid composition comprises anaqueous phase comprising a hydrogel forming polymer, a first surfactantand an oil phase being dispersed in the aqueous phase in whichcyclosporin is dissolved, wherein the first surfactant is or comprises amedium chain or long chain fatty acid mono- or di-glyceride or acombination thereof and the first surfactant does not comprise or is nota polyethyleneglycol ether or ester, the liquid composition, furthercomprising a second surfactant, preferably wherein the second surfactantis an anionic surfactant. The second surfactant may be present in anamount of 10 to 70 mg/g or 15 to 60 mg/g.

Furthermore, in an embodiment the composition comprises cyclosporin, ahydrogel forming polymer matrix, a first surfactant and an oil phasebeing dispersed in the hydrogel forming polymer matrix, wherein thefirst surfactant is or comprises a medium chain or long chain fatty acidmono- or di-glyceride or a combination thereof and does not comprise oris not a polyethyleneglycol ether or ester, the composition furthercomprising a second surfactant, preferably wherein the second surfactantis an anionic surfactant.

Surfactants can also be classified according to theirhydrophilic-lipophilic balance (HLB) which is a measure of the degree towhich the surfactant is hydrophilic or lipophilic, determined bycalculating values for the different regions of the molecule, asdescribed (originally for non-ionic surfactants) by Griffin in 1949 and1954 and later by Davies. The methods apply a formula to the molecularweight of the whole molecule and of the hydrophilic and lipophilicportions to give an arbitrary (semi-empirical) scale up to 40 althoughthe usual range is between 0 and 20. An HLB value of 0 corresponds to acompletely hydrophobic molecule, and a value of 20 would correspond to amolecule made up completely of hydrophilic components. The HLB value canbe used to predict the surfactant properties of a molecule:

HLB Value Expected properties 0 to 3 antifoaming agent from 4 to 6 W/Oemulsifier from 7 to 9 wetting agent from 8 to 18 an O/W emulsifier from13 to 15 typical of detergents 10 to 18 solubiliser or hydrotrope

Although HLB numbers are assigned to surfactants other than thenon-ionic, for which the system was invented, HLB numbers for anionic,cationic, non-ionic, and amphoteric (zwitterionic) surfactants can haveless significance and often represent a relative or comparative numberand not the result of a mathematical calculation. This is why it ispossible to have surfactants above the “maximum” of 20. HLB numbers canhowever be useful to describe the HLB requirement of a desiredapplication for a given emulsion system in order to achieve goodperformance.

Non-Ionic Surfactants

The further surfactant (second surfactant) may be or comprise at leastone surfactant selected from the following non-ionic surfactants.

PEG-fatty acid monoester surfactants, PEG-fatty acid diestersurfactants, PEG-fatty acid monoester and diester surfactant mixtures,PEG glycerol fatty acid esters, transesterified products of oils andalcohols, lower alcohol fatty acid esters, polyglycerised fatty acids,propylene glycol fatty acid esters, mono and diglyceride surfactants,sterol and sterol derivative surfactants, PEG-sorbitan fatty acidesters, sorbitan fatty acid esters, polyethylene glycol alkyl ethers,sugar ester surfactants, polyethylene glycol alkyl phenol surfactants,POE-POP block copolymers, phospholipids.

A PEG-fatty acid mono ester surfactant for example PEG 4-100monolaurate, PEG 4-100 monooleate, PEG 4-100 monostearate, PEG-laurate,PEG-oleate, PEG stearate, and PEG ricinoleate.

A PEG-fatty acid diester surfactant for example PEG dilaurate; PEGdioleate, PEG distearate, PEG dipalmitate. A mixture of PEG-fatty acidmono- and diesters.

A PEG glycerol fatty acid ester for example PEG glyceryl laurate, PEGglyceryl stearate, PEG glyceryl oleate.

PEG-sorbitan fatty acid esters for example PEG sorbitan laurate, PEGsorbitan monolaurate, PEG sorbitan monopalmitate, PEG sorbitanmonostearate, PEG sorbitan tristearate, PEG sorbitan tetrastearate, PEGsorbitan monooleate, PEG sorbitan oleate, PEG sorbitan trioleate, PEGsorbitan tetraoleate, PEG sorbitan monoisostearate, PEG sorbitolhexaoleate, PEG sorbitol hexastearate.

Propylene glycol fatty acid esters for example propylene glycolmonocaprylate, propylene glycol monolaurate, propylene glycol oleate,propylene glycol myristate, propylene glycol monostearate, propyleneglycol 74ydroxyl stearate, propylene glycol ricinoleate, propyleneglycol isostearate, propylene glycol monooleate, propylene glycoldicaprylate/dicaprate, propylene glycol dioctanoate, propylene glyconcaprylate/caprate, propylene glycol dilaurate, propylene glycoldistearate, propylene glycol dicaprylate, propylene glycol dicaprate.

A sorbitan fatty acid ester for example sorbitan monolaurate, sorbitanmonopalmitate, sorbitan monooleate, sorbitan monostearate, sorbitantrioleate, sorbitan sesquioleate, sorbitan tristearate, sorbitanmonoisostearate, sorbitan sesquistearate.

Lower alcohol fatty acid esters for example ethyl oleate, isopropymyristate, isopropyl palmitate, ethyl linoleate, isopropyl linoleate.

Polyoxyethylene-polyoxypropylene block copolymers for example poloxamer105, poloxamer 108, poloxamer 122, poloxamer 123, poloxamer 124,poloxamer 181, poloxamer 182, poloxamer 183, poloxamer 184, poloxamer185, poloxamer 188, poloxamer 212, poloxamer 215, poloxamer 217,poloxamer 231, poloxamer 234, poloxamer 235, poloxamer 237, poloxamer238, poloxamer 282, poloxamer 284, poloxamer 288, poloxamer 331,poloxamer 333, poloxamer 334, poloxamer 335, poloxamer 338, poloxamer401, poloxamer 402, poloxamer 403, poloxamer 407.

Polyglycerised fatty acids for example polyglyceryl stearate,polyglyceryl oleate, polyglyceryl isostearate, polyglyceryl laurate,polyglyceryl ricinoleate, polyglyceryl linoleate, polyglycerylpentaoleate, polyglyceryl dioleate, polyglyceryl distearate,polyglyceryl trioleate, polyglyceryl septaoleate, polyglyceryltetraoleate, polyglyceryl decaisostearate, polyglyceryl decaoleate,polyglyceryl monooleate, dioleate, polyglyceryl polyricinoleate.

PEG alkyl ethers for example PEG oleyl ether, PEG lauryl ether, PEGcetyl ether, PEG stearyl ether.

PEG alkyl phenols for example PEG nonyl phenol, PEG octyl phenol ether.

Transesterification products of alcohol or polyalcohol with natural orhydrogenated oils for example PEG castor oil, PEG hydrogenated castoroil, PEG corn oil, PEG almond oil, PEG apricot kernel oil, PEG oliveoil, PEG-6 peanut oil, PEG hydrogenated palm kernel oil, PEG palm kerneloil, PEG triolein, PEG corn glycerides, PEG almond glycerides, PEGtrioleate, PEG caprylic/capric triglyceride, lauroyl macrogol glyceride,stearoyl macrogol glyceride, mono, di, tri, tetra esters of vegetableoils and sorbitol, pentaerythrityl tetraisostearate, pentaerythrityldistearate, pentaerythrityl tetraoleate, pentaerythrityl tetrastearate,pentaerythrityl tetracaprylate/tetracaprate, pentaerythrityltetraoctanoate.

Oil-soluble vitamins for example vitamins A, D, E, K, and isomers,analogues, and derivatives thereof. The derivatives include, forexample, organic acid esters of these oil-soluble vitamin substances,for example the esters of vitamin E or vitamin A with succinic acid.Derivatives of these vitamins include tocopheryl PEG-1000 succinate(Vitamin E TPGS) and other tocopheryl PEG succinate derivatives withvarious molecular weights of the PEG moiety, for example PEG 100-8000.

Sterols or sterol derivatives (e.g. esterified or etherified sterols asfor example PEGylated sterols) for example cholesterol, sitosterol,lanosterol, PEG cholesterol ether, PEG cholestanol, phytosterol, PEGphytosterol.

Sugar esters for example sucrose distearate, sucrosedistearate/monostearate, sucrose dipalmitate, sucrose monostearate,sucrose monopalmitate, sucrose monolaurate, alkyl glucoside, alkylmaltoside, alkyl maltotrioside, alkyl glycosides, derivatives and othersugar types: glucamides.

Carboxylates (in particular carboxylate esters) for example ethercarboxylates, succinylated monoglycerides, sodium stearyl fumarate,stearoyl propylene glycol hydrogen succinated, mono/diacetylatedtartaric acid esters of mono- and diglycerides, citric acid esters ofmono-, diglycerides, glyceryl-lacto esters of fatty acids; acyllactylates: lactylic esters of fatty acids, calcium/sodiumstearoyl-2-lactylate calcium/sodium stearoyl lactylate, alginate salts,propylene glycol alginate.

A fatty acid monoglyceride, diglyceride or triglyceride or a combinationthereof.

Anionic Surfactants

The further surfactant (second surfactant) may be or comprise at leastone anionic surfactant.

The second surfactant may be a fatty acid salt or bile salt for examplesodium caproate, sodium caprylate, sodium caprate, sodium laurate,sodium myristate, sodium myristolate, sodium palmitate, sodiumpalmitoleate, sodium oleate, sodium ricinoleate, sodium linoleate,sodium linolenate, sodium stearate, sodium lauryl sulfate, sodiumtetradecyl sulfate, sodium lauryl sarcosinate, sodium dioctylsulfosuccinate; sodium cholate, sodium taurocholate, sodiumglycocholate, sodium deoxycholate, sodium taurodeoxycholate, sodiumglycodeoxycholate, sodium ursodeoxycholate, sodium chenodeoxycholate,sodium taurochenodeoxycholate, sodium glyco chenodeoxycholate, sodiumcholylsarcosinate and sodium N-methyl taurocholate. Preferably thesecond surfactant is sodium lauryl sulphate.

Phospholipids for example egg/soy lecithin, cardiolipin, sphingomyelin,phosphatidylcholine, phosphatidyl ethanolamine, phosphatidic acid,phosphatidyl glycerol, phosphatidyl serine.

Phosphoric acid esters having the general formula RO-PO3-M+ where the Rgroup is an ester forming group, e.g. an alkyl, alkenyl or aryl groupoptionally substituted by a PEG moiety through which the alkyl, alkenylor aryl group is coupled to the phosphate moiety. R may be a residue ofa long chain (e.g. >09) alcohol or a phenol. Specific examples includediethanolammonium polyoxyethylene-10 oleyl ether phosphate,esterification products of fatty alcohols or fatty alcohol ethoxylateswith phosphoric acid or anhydride.

Sulfates and sulfonates (in particular esters thereof) for exampleethoxylated alkyl sulfates, alkyl benzene sulfones, α-olefin sulfonates,acyl isethionates, acyl taurates, alkly glyceryl ether sulfonates, octylsulfosuccinate disodium, disodium undecylenamideo-MEA-sulfosuccinate,alkyl phosphates and alkyl ether phosphates.

Cationic Surfactants

The further surfactant (second surfactant) may be or comprise at leastone cationic surfactant selected from the following cationicsurfactants.

Hexadecyl triammonium bromide, dodecyl ammonium chloride, alkylbenzyldimethylammonium salts, diisobutyl phenoxyethoxydimethylbenzylammonium salts, alkylpyridinium salts; betains (trialkylglycine):lauryl betaine (N-lauryl,N,N-dimethylglycine); ethoxylated amines:polyoxyethylene-15 coconut amine, alkyl-amines/diamines/quaternaryamines and alkyl ester.

Emulsifiers

The surfactant may act as an emulsifier such surfactants includenon-ionic emulsifiers, for example selected from: a mixture oftriceteareth-4 phosphate, ethylene glycol palmitostearate and diethyleneglycol palmitostearate (for example sold under the trade mark SEDFOS™75); sorbitan esters, e.g. sorbitan monooleate, sorbitan monolaurate,sorbitan monpalmitate, sorbitan monostearate (for example products soldunder the trade mark Span®), PEG-8 beeswax e.g. sold under the trademark Apifil®; a mixture of cetyl alcohol, ceteth-20 and steareth-20 (forexample Emulcire™ 61 WL 2659); a mixture of PEG-6 stearate and PEG-32stearate (for example Tefose® 1500); a mixture of PEG-6 palmitostearate,ethylene glycol palmitostearate, and PEG-32 palmitostearate (e.g.Tefose® 63); triglycerol diisostearate (for example products sold underthe trade mark Plurol Diisostearique®; polyglyceryl-3 dioleate (forexample products sold under the trade mark Plurol® Oleique).

Preferred Second Surfactants.

Preferably the second surfactant is an anionic surfactant. For example,the second surfactant may be an alkyl sulphate, for example sodiumdodecyl sulphate. The second surfactant may be present in an amount of10 to 70 mg/g or 15 to 60 mg/g.

The second surfactant may be a fatty acid salt or bile salt for examplesodium caproate, sodium caprylate, sodium caprate, sodium laurate,sodium myristate, sodium myristolate, sodium palmitate, sodiumpalmitoleate, sodium oleate, sodium ricinoleate, sodium linoleate,sodium linolenate, sodium stearate, sodium lauryl sulfate, sodiumtetradecyl sulfate, sodium lauryl sarcosinate, sodium dioctylsulfosuccinate; sodium cholate, sodium taurocholate, sodiumglycocholate, sodium deoxycholate, sodium taurodeoxycholate, sodiumglycodeoxycholate, sodium ursodeoxycholate, sodium chenodeoxycholate,sodium taurochenodeoxycholate, sodium glyco chenodeoxycholate, sodiumcholylsarcosinate and sodium N-methyl taurocholate. Preferably thesecond surfactant is sodium lauryl sulphate.

Other Excipients

The composition optionally contains one or more of the followingadditional substances or categories of substances. For example, thecomposition may contain a protectant such as, for example, a proteolyticenzyme inhibitor or a protector against acid degradation or both (e.g.an alkali for example sodium hydroxide); an adhesive entity such as, forexample, a muco- or bio-adhesive; excipients to maximize solubility ofthe active ingredient; excipients to maximize permeability of the activeingredient in the GIT. Typical excipients for enhancing the permeabilityof the epithelial barrier include but are not limited to sodium caprate,sodium dodecanoate, sodium palmitate, SNAC, chitosan and derivativesthereof, fatty acids, fatty acid esters, polyethers, bile salts,phospholipids, alkyl polyglucosides, sugar esters, hydroxylaseinhibitors, antioxidants (e.g. ascorbic acid) and/or nitric oxidedonors. The preceding list is of particular interest to enhancepermeability in the ileum.

To enhance permeability in the colon, typical excipients include, butnot limited to sodium caprate, sodium dodecanoate, sodium palmitate,SNAC, chitosan and derivatives thereof, fatty acids, fatty acid esters,polyethers, bile salts, phospholipids, alkyl polyglucosides, hydroxylaseinhibitors, antioxidants (optionally selected from curcuminoids,flavonoids, curcumin, beta-carotene, α-tocopherol, ascorbic acid,ascorbate, lazaroid, carvedilol, butylated hydroxytoluene, propylgallate, hydralazine, carnosic acid, vitamin E, lecithin Ovolecithin(vitelin), vegilecithin, fumaric acid or citric acid) and/or nitricoxide donors, including nitric oxide donor groups covalently attached tovarious pharmaceutically active ingredients. The composition may furthercomprise excipients or other active pharmaceutical or other ingredientsto enhance local tissue bioavailability in the GIT, such as the smallintestine or colon, including efflux pump inhibitors, including, but notlimited to PgP pump inhibitors (optionally selected from NSAIDs,cimetidine, omeprazole, Vitamin E TPGS, verapimil, quinidine, PSC833,amprenavir (APV), indinavir (IDV), nelfinavir (NFV), ritonavir (RTV) andsaquinavir (SQV)), and metabolism inhibitors, including, but not limitedto, cytochrome P450 inhibitors, optionally selected from: essentialoils, cimetidine, surfactants (for example cremophor), oils, omeprazole,verapamil, ritonavir and carbamazepine as well as plant extracts, e.g,from citrus fruits. The composition may therefore further comprise aP450 inhibitor to further reduce metabolism of cyclosporin followingadministration of the composition. The P450 inhibitor may act to inhibitenteric and/or hepatic metabolism of the cyclosporin. The compositionmay further comprise a PgP inhibitor. Optionally the composition maycomprise a P450 inhibitor and a PgP inhibitor.

The composition may further comprise excipients to enhance thetherapeutic potential of an active ingredient, for example cyclosporin Aor another immunosuppressant, throughout the gastrointestinal tract,including in the ileum and colon including, but not limited toabsorption limiters, essential oils such as, for example, omega 3 oils,natural plant extracts such as, for example, neem, ion-exchange resins,bacteria degradable conjugation linkers such as, for example, azo bonds,polysaccharides such as, for example, amylose, guar gum, pectin,chitosan, inulin, cyclodextrins, chondroitin sulphate, dextrans, guargum and locust bean gum, nuclear factor kappa B inhibitors, acids suchas, for example, fumaric acid, citric acid and others, as well asmodifications thereof.

The composition may further comprise excipients to reduce systemic sideeffects associated with absorption of certain active, for examplecyclosporin or other immunosuppressants, in the GIT, such as the smallintestine, including, but not limited to, antioxidants, such as, forexample, curcuminoids, flavanoids or more specifically includingcurcumin, beta-carotene, α-tocopherol, ascorbate or lazaroid.

The composition may further or separately comprise antioxidants (suchas, for example, ascorbic acid or BHT—butyl hydroxy toluene)taste-masking or photosensitive components or photoprotectivecomponents. Antioxidants may be incorporated in the aqueous phase (e.g.hydrophilic antioxidants) or in the disperse phase of the core (e.g.hydrophobic antioxidants such as, for example, vitamin E) for example upto 1% by weight, preferably between 0.01 and 0.50% by weight, morepreferably between 0.10 to 0.20% by weight.

The composition may further comprise immune-enhancing nutrients such asvitamins A/B/C/E; carotenoids/beta-carotene and iron, manganese,selenium, zinc, especially when the composition contains animmunosuppressant, as in the case of an immunosuppressant targeted tothe ileum and/or colon, e.g. the colon. Such nutrients may be present incomposition, or if the composition has a coating, for example if it isthe form of a bead, the nutrients may be included in the coating.

The composition may also include other well know excipients used inpharmaceutical compositions including colorants, taste masking agents,diluents, fillers, binders etc. The presence of such optional additionalcomponents will of course depend upon the particular dosage formadopted.

Shape, Size and Geometry

The composition of the invention can be formed into a limitless numberof shapes and sizes. In the section below describing the process formaking the composition, various methods are given including pouring orintroducing a fluid dispersion into a mould where it hardens or can becaused to harden. Thus the composition can be created in whichever formis desired by creating an appropriate mould (e.g. in the shape of adisc, pill or tablet). However, it is not essential to use a mould. Forexample, the composition may be formed into a sheet e.g. resulting frompouring a fluid dispersion onto a flat surface where it hardens or canbe caused to harden.

Preferably, the composition may be in the form of spheres orspherical-like shapes made as described below. Preferably, thecomposition of the invention is in the form of substantially spherical,seamless minibeads. The absence of seams on the minibead surface is anadvantage e.g. in further processing, for example coating, since itallows more consistent coating, flowability etc. The absence of seams onthe minbeads also enhances consistency of dissolution of the beads.

The preferred size or diameter range of minibeads according to theinvention can be chosen to avoid retention in the stomach upon oraladministration of the minibeads. Larger dosage forms are retained forvariable periods in the stomach and pass the pyloric sphincter only withfood whereas smaller particles pass the pylorus independently of food.Selection of the appropriate size range (see below) thus makes thetherapeutic effect post-dosing more consistent. Compared to a singlelarge monolithic oral format such as, for example, a traditionalcompressed pill, a population of beads released into the GI tract (asforeseen by the dosage form of the present invention) permits greaterintestinal lumen dispersion so enhancing absorption via exposure togreater epithelial area, and achieves greater topical coating in certainparts of the GI tract for example the colon). Reduction of residencetime in the ileo-caecal junction is another potential advantage.

The composition of the invention is preferably monolithic meaninginternally (i.e. cross-sectionally) homogeneous, excluding a possiblethin skin of matrix material and excluding any coating layers.

The minibeads provided for by the formulation of the present inventiongenerally range in diameter from 0.5 mm to 10 mm with the upper limitpreferably 5 mm, e.g. 2.5 mm A particularly convenient upper limit is 2mm or 1.7 mm. The lower limit can preferably be 1 mm, e.g. 1.2 mm, morepreferably from 1.3 mm, most preferably from 1.4 mm. In one embodimentthe diameter is from 0.5 to 2.5 mm, for example from 1 mm to 3 mm, 1 mmto 2 mm, 1.2 mm to 3 mm or 1.2 mm to 2 mm. The minibeads may have adiameter of no more than 2.5 mm, irrespective of their minimum size. Thebeads may have a diameter of no more than 2 mm, irrespective of theirminimum size.

A minibead as described herein may have an aspect ratio of no more than1.5, e.g. of no more than 1.3, for example of no more than 1.2 and, inparticular, of from 1.1 to 1.5, 1.1 to 1.3 or, 1.1 to 1.2. A populationof minibeads as described herein, e.g. at least 10 beads, may have anaverage aspect ratio of no more than 1.5, e.g. of no more than 1.3, forexample of no more than 1.2 and, in particular, of from 1 to 1.5, 1 to1.3 or 1 to 1.2. The aspect ratios mentioned in this paragraphoptionally apply to coated minibeads and optionally apply to uncoatedminibeads. Average aspect ratio is suitably determined for a populationof minibeads, e.g. at least 10 minibeads, using a particle sizeanalyser, for example an Eyecon™ particle characteriser of InnopharmaLabs, Dublin 18, Ireland.

The minibeads of the disclosure may, therefore, have a size as disclosedabove and an aspect ratio of from 1 to 1.5. The beads of the disclosuremay have a size as disclosed above and an aspect ratio of no more than1.3, for example of no more than 1.2 and, in particular, of from 1.1 to1.5, 1.1 to 1.3 or, 1.1 to 1.2.

Bead size (diameter) may be measured by any suitable technique, forexample microscopy, sieving, sedimentation, optical sensing zone method,electrical sensing zone method or laser light scattering. For thepurposes of this specification, bead size is measured by analyticalsieving in accordance with USP General Test <786> Method I (USP 24-NF18, (U.S. Pharmacopeial Convention, Rockville, Md., 2000), pp.1965-1967).

In embodiments, minibeads of the invention are monodisperse. In otherembodiments, minibeads of the invention are not monodisperse. By“monodisperse” is meant that for a population of beads (e. g. at least100, more preferably at least 1000) the minibeads have a coefficient ofvariation (CV) of their diameters of 35% or less, optionally 25% orless, for example 15% or less, such as e.g. of 10% or less andoptionally of 8% or less, e.g. 5% or less. A particular class of polymerbeads has a CV of 25% or less. CV when referred to in this specificationis defined as 100 times (standard deviation) divided by average where“average” is mean particle diameter and standard deviation is standarddeviation in particle size. Such a determination of CV is performableusing a sieve.

The invention includes minibeads having a CV of 35% and a mean diameterof 1 mm to 2 mm, e.g. 1.5 mm. The invention also includes minibeadshaving a CV of 20% and a mean diameter of 1 mm to 2 mm, e.g. 1.5 mm, aswell as minibeads having a CV of 10% and a mean diameter of 1 mm to 2mm, e.g. 1.5 mm. In one class of embodiments, 90% of minibeads have adiameter of from 0.5 mm to 2.5 mm, e.g. of from 1 mm to 2 mm.

Dosage Forms

The composition of the invention may be prepared as an orallyadministrable dosage form suitable for pharmaceutical use. In thoseembodiments where the formulation is in the form of a minibead, thepresent invention provides for a dosage form comprising a plurality ofthe minibeads for example as a capsule, a tablet, a sprinkle or asachet. The minibeads may also be administered rectally or vaginallyadministered composition, for example as an enema or suppository. Thecomposition, for example in the form of minibeads may be blended in asuitable medium to provide a suppository or enema compostions. Suitablemedia for suppositories and enemas are well known and include forexample, a low melting point wax for a suppository or a suitable aqueousor oil based medium for an enema composition.

The liquid composition of the invention may be formulated as an orally,rectally or vaginally administrable dosage form suitable forpharmaceutical use. The liquid composition may be formulated into a hardor soft gelatin capsule, a suppository or an enema. Delivery of theliquid composition to the stomach may also be achieved via a gastricfeeding tube located in the stomach or by means of a percutaneousendoscopic gastrostomy tube (PEG tubing) as hereinabove described. Theliquid composition may also be administered directly to specific pointsin the GI tract, for example the duodenum, jejunem or ileum via oral orintranasal tubing with an exit at the desired point in the GI tract.Delivery of the liquid composition via tubing may be under gravity flowor under positive pressure using a pump or syringe drive etc.

In embodiments the dosage form comprising a population of beads may bepresented in a single unit dosage form e.g. contained in a single hardgel capsule which releases the beads e.g. in the stomach. Alternativelythe beads may be presented in a sachet or other container which permitsthe beads to be sprinkled onto food or into a drink or to beadministered via a feeding tube for example a naso-gastric tube or aduodenal feeding tube. Alternatively, the beads may be administered as atablet for example if a population of beads is compressed into a singletablet as described below. Alternatively, the beads may be filled e.g.compressed into a specialist bottle cap or otherwise fill a space in aspecialised bottle cap or other element of a sealed container (orcontainer to be sealed) such that e.g. on twisting the bottle cap, thebeads are released into a fluid or other contents of the bottle or vialsuch that the beads are disperse (or dissolve) with or without agitationin such contents. The fluid or other contents of the bottle or vial mayoptionally contain one of more additional active agent(s) to facilitatethe conventiaent co-administration of the cyclosporin composition withother active agents. An example is the Smart Delivery Cap manufacturedby Humana Pharma International (HPI) S.p.A, Milan, Italy.

The dosage form may be formulated in such a way so that the beads of theinvention can be further developed to create a larger mass of beads e.g.via compression (with appropriate oil or powder-based binder and/orfiller known to persons skilled in the art. The larger (e.g. compressed)mass may itself take a variety of shapes including pill shapes, tabletshapes, capsule shapes etc. A particular problem which this version ofthe bead embodiment solves is the “dead space” (above the settledparticulate contents) and/or “void space” (between the particulatecontent elements) typically found in hard gel capsules filled withpowders or pellets. In such pellet- or powder-filled capsules withdead/void space, a patient is required to swallow a larger capsule thanwould be necessary if the capsules contained no such dead space. Thebeads of this embodiment of the invention may readily be compressed intoa capsule to adopt the inner form of whichever capsule or shell may bedesired leaving much reduced, e.g. essentially no, dead/void space.Alternatively the dead or void space can be used to advantage bysuspending beads in a vehicle such as, for example, an oil which may beinert or may have functional properties such as, for example,permeability enhancement or enhanced dissolution or may comprise anactive ingredient being the same or different from any activeingredients in the bead. For example, hard gelatin or HPMC capsules maybe filled with a liquid medium combined with uncoated and/or coatedbeads. The liquid medium may be one or more of the surfactant phaseconstituents described herein or it may be one or more surfactants.Particularly preferred but non-limiting examples are corn oil, sorbitanetrioleate (sold under the trade mark SPAN 85), propylene glycoldicaprylocaprate (sold under the trade mark Labrafac),2-(2-ethoxyethoxy)ethanol (sold under the trade mark Transcutol P) andpolysorbate 80 (sold under the trade mark Tween 80).

In a representative embodiment the bead of the dosage form is preparedas described herein for example by mixing together at least thefollowing materials: a hydrogel-forming polymer; an oil phase, asurfactant being or comprising a medium chain or long chain fatty acidmono- or di-glyceride or a combination thereof, wherein the surfactantdoes not comprise or is not a polyethyleneglycol ether or ester, andcyclosporin A, suitably cyclosporin A being dissolved in the oil phase,such as a liquid lipid to form a dispersion of the cyclosporin A in thehydrogel-forming polymer. The dispersion is immobilized within thesolidified bead by ejection from a single orifice nozzle into a suitablecooling liquid. Following removal of the cooling liquid the bead iscoated with a modified release coating (the second coating) (suitablywith a sub-coat under the modified release coating), the coated bead isthen optionally filled into a gelatin or HPMC capsule suitable forpharmaceutical use.

Suitably the dosage form is prepared as a unit dosage form containingfrom for oral administration comprising from 0.1 mg to 1000 mg,optionally from 1 mg to 500 mg, for example 10 mg to 300 mg, 15 mg to300 mg, or 25 to 250 mg, suitably about 15 mg, about 25 mg, about 35 mg,about 50 mg, about 75 mg, about 100 mg, about 150 mg, about 180 mg,about 200 mg, about 210 mg or about 250 mg cyclosporin A

Determination of Contents and Distribution of Formulations

The identity and/or distribution of one or more of the components of acomposition according to the invention can be determined by any methodknown to those skilled in the art. The distribution of one or morecomponents of a composition can, for example, be determined bynear-infrared (NIR) chemical imaging technology. NIR chemical imagingtechnology can be used to generate images of the surface or crosssection of a composition, for example a minibead. The image produced bythis technique shows the distribution of one or more components of thecomposition. In addition to NIR chemical imaging technology, thedistribution of one or more components of a composition such asminibead, for example, be determined by time-of-flight secondary ionmass spectrometry (ToFSIMS). ToFSIMS imaging can reveal the distributionof one or more components within the composition. The images produced byToFSIMS analysis or NIR analysis can show the distribution of componentsacross a surface of the composition or a cross section of thecomposition. The methods described in this paragraph are applicable, forexample, to composition comprising a polymer matrix, e.g. a dried,colloid, solution or dispersion.

Pharmacokinetics

The orally administered compositions comprising cyclosporin A describedherein may provide, amongst other features, a favourable pharmacokineticprofile compared to orally administered Neoral and/or to intravenouslyadministered cyclosporin A as for example Sandimmun™.

The compositions according to the invention provide lower mean wholeblood exposure to cyclosporin A following oral administration comparedto oral administration of Neoral™ at the same dose of cyclosporin A. Thewhole blood exposure to cyclosporin A may be determined by measuring thearea under the Curve (AUC) of the whole blood cyclosporin Aconcentration-time curve following administration of a single dose of acomposition containing cyclosporin A. The area under theconcentration-time curve (AUC), calculated from the start of dosing(t=0) to the last measured concentration (t) is designated to be“AUC_(0-t)”. Accordingly reference to “AUC₀₋₂₄ hr” is the AUC betweent=0 and the last measurement point at 24 hours following administration.The AUC_(0-t) may be calculated using well known methods for example bylinear trapezoidal analysis. The area under the concentration-time curveextrapolated to infinity is “AUC_(0-inf)”. The AUC_(0-inf) is calculatedusing known methods as:

${AUC}_{0 - t} + \frac{C_{t}}{K_{el}}$

Where: C_(t)=the fitted last non-zero concentration for that treatment,AUC_(0-t) is as defined above; and K_(el)=the elimination rate constant.K_(el) is calculated by regression analysis of the natural log (Ln) ofwhole blood concentration values-time profile.

The term “Cmax” refers to the maximum concentration of cyclosporin inwhole blood following administration of a single dose of a compositioncontaining cyclosporin A.

The term “Tmax” refers to the time taken to reach Cmax following oraladministration of a composition containing cyclosporin A.

For statistical analysis, the PK data is log-transformed prior toconducting statistical testing. In general, statistical tests arecarried out using an analysis of variance procedure (ANOVA) andcalculating a 90% confidence interval for each pharmacokinetic parameter(Cmax and AUC).

The measurement and analysis of AUC, Cmax and Tmax are well known in theart and can be carried out using methods and techniques described infurther detail in the examples or by reference to standard textbookssuch as Remington, The Science and Practice of Pharmacy 22^(nd) edition,or Basic Pharmacokinetics and Pharmacodynamics: An integrated Textbookand Computer Simulations, Sara E. Rosenbaum, 2011 John Wiley& Sons. Inall cases references to AUC, Cmax and Tmax are the mean values measuredfollowing administration of a composition containing cyclosporin A to ahuman, preferably a healthy male human in a fasted state. Suitably thesubjects used in the PK study are adult male humans with weighing about70 kg (for example 70 kg±12 kg). Suitably the subjects have a body massindex of about 25 kg/m² (for example 25 kg/m²±2.5 kg/m²).

In some embodiments the composition of the invention provides an AUCand/or a Cmax value as the AUC or Cmax “following oral administration ofa single dose of 75 mg cyclosporin A”. It is known that cyclosporin Aexhibits an approximately linear pharmacokinetic profile EU HMA's PublicAssessment Report on Ciclosporin “Docpharma” soft capsulesDK/H/968/1-3/MR, page 4.

Accordingly reference to an “AUC or Cmax of a particular value afteroral administration of the composition as a single dose containing 75 mgcyclosporin A to a human in a fasted state, or an AUC or Cmax directlyproportional thereto for a total dose other than 75 mg” is to beunderstood to mean that the AUC or Cmax value is directly proportionalto mass of the cyclosporin A dose administered. By way of example, if asingle dose of 150 mg cyclosporin A were to be administered thecorresponding AUC and Cmax values will be approximately twice thatobtained with a single dose of 75 mg cyclosporin A. Similarlyadministration of a single dose of 37.5 mg of cyclosporin A would beexpected to provide a AUC and Cmax values approximately half thoseobserved following administration of 75 mg cyclosporin A. The doseproportionality for cyclosporin A is applicable over a broad range ofdosages of cyclosporin A for example from 0.1 to 1000 mg, suitablybetween about 1 mg and about 500 mg, more particularly between about 5mg and about 350 mg.

In one embodiment the composition provides a mean whole bloodAUC_(0-inf) of from about 140 to about 420 ng·hr/ml, for example fromabout 140 to about 350 ng·hr/ml, about 140 to about 400 ng·hr/ml about150 to about 350 ng·hr/ml, about 150 to about 300 ng·hr/ml about 180 toabout 350 ng·hr/ml, about 200 to about 400 ng·hr/ml or about 180 toabout 320 ng·hr/ml and a mean whole blood Cmax of from about 25 to about45 ng/ml after oral administration of the composition as a single dosecontaining 75 mg cyclosporin A to a human in a fasted state, or aAUC_(0-inf) and Cmax directly proportional thereto for a total doseother than 75 mg. Suitably the composition provides a Tmax of from about4 hours to about 8 hours, suitably from about 4 hours to about 6 hoursand particularly at about 5 hours; or about 5.5 hours; or about 6 hours.

Cyclosporin A Concentration in Faecal Samples

The cyclosporin composition may release cyclosporin A (preferably in asolubilised form for example as a solution in an oil droplet or asmicelles containing cyclosporin A) in the lower GI tract andparticularly the colon. Accordingly, the composition provides high localcyclosporin A concentrations in the luminal contents and further resultsin absorption of cyclosporin A into the tissue of the GI tract. Theluminal and tissue concentration of cyclosporin A following oraladministration of a composition of the invention is higher relative tothat resulting from oral administration of Neoral™ or IV administrationas Sandimmun™. However, as discussed above, the composition according tothe invention results in a relatively low systemic blood exposure to thecyclosporin A. The cyclosporin compositions described herein comprisingthe surfactant may also reduce the cyclosporin A metabolism followingrelease of the cyclosporin from the composition into the GI tract.

The cyclosporin composition may provide a ratio of the meanconcentration of cyclosporin A:the concentration of cyclosporin Ametabolites (for example the sum of the mean AM4N and AM9 metaboliteconcentrations, or the sum of the mean AM1, AM9, and AM4N metaboliteconcentrations of greater than 12:1. Suitably the metaboliteconcentration is measured as the sum of the mean concentration of eachmetabolite present in the faecal sample. In one embodiment “theconcentration of cyclosporin A metabolites” refers to the sum of themean concentrations of the AM4N+AM9 metabolites present in the sample.In another embodiment “the concentration of cyclosporin A metabolites”refers to the sum of the mean concentrations of the AM4N+AM9+AM1metabolites present in the sample. Accordingly In one embodiment ratioof the mean concentration of cyclosporin A:the concentration of AM1,AM9, and AM4N metabolites is greater than 12:1, for example, from 20:1to 40:1, from 20:1 to 35:1, Suitably the ratio of cyclosporinA:metabolite concentration in the faecal sample is determined afterorally administering a single dose of 75 mg cyclosporin A. However,other doses and dose regimens such as twice daily dosing may also beused. As described above the concentrations may be determined in afaecal sample collected 12 to 28 hours after dosing the composition.However, the concentrations may be determined in faeces collected atother time points following oral administration of the compositionprovided sufficient time has elapsed after oral administration of thecomposition for transit through the gut such that cyclosporin and itsmetabolites to be present in the collected faecal sample. It is expectedthat the ratios of cyclosporin to metabolites measured in a collectedfaecal samples will be approximately the same irrespective of thespecific time point at which the faeces is collected. Accordingly,reference herein to collection of a faecal sample at 12 to 28 hours isnot intended to be limiting. Suitably, the ratios of cyclosporin tometabolites are measured in samples of faeces taken from subjects thathave been exposed to a regular daily dose of the composition. After aprolonged period of daily dosing it is expected that steady-stateconcentrations of cyclosporin and metabolites will be achieved and assuch there may be less variability in the measured concentrations ofcyclosporin and metabolites in the faeces. Accordingly, theconcentration of cyclosporin: metabolites may, for example, be measuredin a faecal sample collected 4 to 6 hours after oral administration ofthe last dose of a once daily oral dosing regimen of the composition,the dosing regimen comprising once daily oral administration of thecomposition (for example containing 75 mg cyclosporin A) for seven days.

By way of comparison to the compositions according to the invention, theexamples herein show that oral administration of Neoral™ results in aratio of cyclosporin A:Cyclosporin metabolites (AM4N+AM9) ofapproximately 0.6:1, reflecting the relatively high systemic exposureand relatively low local tissue exposure in the lower GI tract,particularly in the colon. Similarly IV administration of 2 mg/Kg ofcyclosporin resulted in a ratio of about 0.3:1 to about 0.45:1.

Cyclosporin A in Luminal Contents and GI Tissue

The high concentration of cyclosporin A in the luminal contents of thelower GI tract and the concentration of cyclosporin A in the tissue ofthe GI tract may be determined by measuring cyclosporin A concentrationin luminal content and tissue samples taken at specific points along theGI tract. Cyclosporin A concentrations in intracolonic faeces andcolonic tissue may be measured in human patients as described in theprotocols described in the Examples. The composition comprisingcyclosporin provides high concentrations of cyclosporin A in the mucosaand sub-mucosa (i.e. the inner tissues) of for example the colon. Thecyclosporin A concentration in the colonic tissues may be measured bytaking a section of the colonic tissue, separating the layers of tissue(for example the mucosa, sub-mucosa and muscularis externa), andmeasuring the cyclosporin concentration in each of the respective tissuelayers.

The presence of a high colonic luminal cyclosporin A concentrationprovided by the composition of the invention is expected to provide aconcentration gradient which acts to promote absorption of thecyclosporin A (preferably in a solubilised form) into the lamina propriaof the colon, where the main target dysregulated immune cells associatedwith many inflammatory diseases of the colon predominate. Thecompositions of the invention therefore provide a local topicaltreatment of diseased colonic tissue and are expected to be useful inthe treatment of conditions such as ulcerative colitis and otherinflammatory diseases affecting the at least the colon. In contrast oraladministration of Neoral™ provides relatively low luminal concentrationof cyclosporin A to the inner colonic tissues. Intravenousadministration of cyclosporin A as Sandimmun™ reduces the metabolism inthe intestine and may provide similar faecal metabolite concentrationsas an orally administered composition according to the invention.However, the IV administration of cyclosporin results in significantlyhigher systemic exposure and moreover, relatively high doses of IVcyclosporin may be required to provide therapeutic concentrations ofcyclosporin in the colonic tissue compared to oral administration of acomposition according to the invention.

The composition comprising cyclosporin may therefore be expected toprovide a therapeutic benefit at lower doses than Neoral and/orSandimmun™, thus further minimising side effects associated withsystemic exposure to cyclosporin A. Some release or cyclosporin A fromthe composition may occur as the composition passes through the GI tractand release of cyclosporin may not be exclusive to the colon. As suchthe composition of the invention may provide locally acting cyclosporinA in at least the colon and in other parts of the GI tract, for examplethe rectum and ileum, the composition may therefore provide therapeuticbenefit in the treatment or prevention of conditions affecting not justthe colon, but also other parts of the GI tract as described herein.

Measurement of cyclosporin concentration in the colonic tissue andintracolonic faeces in humans may be performed as described in theExamples. Suitably samples of colonic tissue are obtained from a patientwho has been orally treated with a composition containing cyclosporin bysigmoidoscopy using, for example pinch biopsy forceps, to obtain samplesof colonic tissue. Suitably sigmoidoscopy is a flexible sigmoidoscopy.The sigmoidoscopy is preferably carried out in the unprepared bowel(except for air and water) such that the tissue samples obtainedreplicate as closely as possible the in-vivo tissue status, which mightotherwise be disturbed by extensive bowel preparation. Biopsies aresuitably about 5 mm in size and ideally at least 5 biopsies are takenapproximately 1 cm apart from the subject. Preferably the biopsies areobtained as close to the splenic flexure as possible. Alternatively,biopsies may also be obtained from within the sigmoid colon. Each biopsyshould be rinsed with saline, blot dried and then stored at lowtemperature, suitably at about −70° C., prior to analysis. The tissuesamples may be analysed directly for the concentration of cyclosporin Apresent in the tissue. However, preferably the mucous layer present onthe tissue surface is first removed from the sample such that thecyclosporin concentration measured is the concentration of cyclosporinpresent in the epithelial and musosal tissue. The mucous layer may beremoved by washing with a suitable solvent such as N-acetyl cysteine inwater. Removal of the mucose layer ensures that the concentration ofcyclosporin measured is representative of the concentration which hasbeen absorbed into the tissue, rather than the mucosal layer. Thecyclosporin present in the mucosal layer can be determined by analysingthe mucosal washings. Suitable methods for the preparation and analysisof the colonic tissue are set out in the examples section.

Samples of intracolonic faeces are suitably collected from approximatelythe same location within the colon as the tissue biopsies such that themeasurement of cyclosporin concentration in the tissue and intra-colonicfaeces represents the concentrations present at approximately the sameposition within the colon.

The tissue biopsies and intra-colonic faecal samples should be obtainedafter a sufficient duration of cyclosporin dosing to reach steady-stateconcentrations in the colon. For example, the biopsies and faecalsamples are suitably may be carried out after 7 days of daily oraldosing with the composition. The biopsies and intracolonic faecalsamples are suitably obtained simultaneously within 4 to 6 hours afterthe last dose in the 7 day dosage regimen.

The ratio of the mean concentration of cyclosporin A present inintracolonic faeces:the mean concentration of cyclosporin A present incolonic tissue in an adult human patient after oral administration ofthe composition is greater than 30:1, for example, greater than about40:1 or greater than about 50:1. The mean concentration of cyclosporin Apresent in intracolonic faeces:the mean concentration of cyclosporin Apresent in colonic tissue may be about 30:1 to about 500:1, about 50:1to about 500:1, optionally from about 80:1 to about 300:1, or optionallyabout 100:1 to about 250:1. In contrast the Examples show that IVadministration of Sandimmun results in an intracolonic faecal:tissueratio of cyclosporin A of about 3:1.

References to a “mean” value in relation of the PK, tissue and faecalanalysis described herein is unless specified otherwise a reference tothe arithmetic mean value of the measured values.

Dissolution Profile

The compositions comprising cyclosporin provide compositions with aspecific in-vitro dissolution profile for the release cyclosporin A fromthe composition. The compositions show minimal release of cyclosporin Ain the stomach and upper GI tract such as the duodenum and jejunum andhigher release in at least the colon. The in-vivo release may bemodelled using a two stage in-vitro dissolution test in which acomposition is exposed to 0.1 N HCl for two hours to simulate pH of thegastric environment and is then exposed to pH 6.8 for twenty two hours(by adding a sufficient quantity of 0.2M tribasic sodium phosphatesolution containing 2% sodium dodecyl sulfate (SDS)) to simulate pH inthe small intestine and lower GI tract.

Reference to “a two stage dissolution test using a USP Apparatus II witha paddle speed of 75 rpm and a dissolution medium temperature of 37° C.;wherein for the first 2 hours of the dissolution test the dissolutionmedium is 750 ml of 0.1 N HCl, and at 2 hours 250 ml of 0.2M tribasicsodium phosphate containing 2% SDS is added to the dissolution mediumand the pH is adjusted to pH 6.8” is an in-vitro test carried out inaccordance with the USP <711> Dissolution test using Apparatus II(paddle apparatus) operated with a paddle speed of 75 rpm and with thedissolution medium at a temperature of 37° C.±0.5° C. At the start ofthe test (t=0) the sample is placed in the acidic dissolution medium.After 2 hours an aliquot of the medium is taken for subsequent analysisand immediately (suitably within 5 minutes) the second stage of thedissolution test is initiated. In the second stage of the dissolutiontest 250 ml of 0.2M tribasic sodium phosphate containing 2% sodiumdodecyl sulfate (SDS) is added to the dissolution medium and the pH isadjusted to 6.8±0.05 using 2N NaOH or 2N HCl as required. Samples of thedissolution medium are taken at time points during the second stage ofthe test, for example at 4, 6, 12 and 24 hours from the start of thetest (i.e. from t=0 at the start of the first stage). The samples areanalysed for cyclosporin A dissolved in the medium. The “% released” isthe amount of cyclosporin A in solution in the respective dissolutionmedium at a particular time point relative to the amount of cyclosporinin the composition at the start of the test. The cyclosporin Aconcentrations in a sample may be measured using standard techniques,such as Reverse Phase HPLC as illustrated in the Examples. References to“two stage dissolution test” herein also refer to this test method.

The in-vitro dissolution profile of the composition according to theinvention is described above under the Brief Summary.

Manufacturing Processes

Various methods may be used to prepare the formulations of theinvention.

In those embodiments where the formulation comprises an activeingredient in a water-insoluble polymer matrix, a basic method formaking the core is to mix a fluid form of the matrix material, forexample a hydrogel forming polymer matrix material (e.g. poly(amides),poly(amino-acids), hyaluronic acid; lipoproteins; poly(esters),poly(orthoesters), poly(urethanes) or poly(acrylamides), poly(glycolicacid), poly(lactic acid) and corresponding co-polymers(poly(lactide-co-glycolide acid; PLGA); siloxane, polysiloxane;dimethylsiloxane/methylvinylsiloxane copolymer;poly(dimethylsiloxane/methylvinylsiloxane/methylhydrogensiloxane)dimethylvinyl or trimethyl copolymer; silicone polymers; alkyl silicone;silica, aluminium silicate, calcium silicate, aluminium magnesiumsilicate, magnesium silicate, diatomaceous silica etc as described moregenerally elsewhere herein), with an active ingredient to form a mixturethat may take the form of a suspension, solution or a colloid. Themixture is processed to form a composition or a core. For example thecomposition may be shaped into the desired form using a molding orhot-melt extrusion process to form beads.

Methods for preparing a composition and a core comprising thesurfactant, cyclosporin, an oil phase and a water-soluble polymer matrixare described below. Generally these cores are coated.

Generally, the manufacturing processes described herein comprise mixingof liquid(s). Such mixing processes must be performed at temperatures atwhich the substances to be mixed in the liquid state are in liquid form.For example, thermoreversible gelling agents must be mixed at atemperature where they are in the liquid state, for example at atemperature of 50 to 75° C., for example 50 to 70° C., or 55-75° C.,e.g. 60-70° C. and in particular embodiments about 55° C. or 65° C. inthe case of mixing formulations comprising aqueous gelatin. Similarlyother components of the formulation may need to be heated to melt thecomponent for example waxes or surfactants which may be used in thedisperse phase.

The liquid composition, composition or the core comprising a surfactant,oil phase, hydrogel-forming polymer and cyclosporin as disclosed hereinmay be made by mixing materials comprising for example water, ahydrogel-forming polymer and a second surfactant to form an aqueouscontinuous phase, and mixing a disperse phase. At least one of theaqueous phase and the disperse phase comprises cyclosporin, thecyclosporin may be dissolved in the phase which contains it, for exampleboth phases may be a clear liquid before they are mixed together.Preferably, the disperse phase (the oil phase) may comprise thecyclosporin, (for example a disperse phase comprising an oil, anoptional solvent, cyclosporin and a first surfactant) with the aqueousphase to form a colloid. The colloid may have the form of an emulsion ormicroemulsion wherein the disperse phase is dispersed in the aqueouscontinuous phase. This colloid may optionally represent the liquidcomposition of the invention. In order to prepare the composition of theinvention or the core, the hydrogel-forming polymer is then caused orallowed to gel to form a hydrogel forming polymer matrix. Suitably, theprocess includes formulating or processing the composition into adesired form, e.g. a bead (also termed a minibead), which formingprocess may comprise moulding but preferably comprises ejecting theaqueous colloid through a single orifice nozzle to form droplets whichare caused or allowed to pass into a cooling medium, e.g. awater-immiscible cooling liquid, in which the droplets cool to form fore.g. beads.

The mixing of the materials may comprise mixing an aqueous premix (oraqueous phase or continuous phase) and a disperse phase premix (e.g. oilphase premix), wherein the aqueous premix comprises water andwater-soluble substances whilst the disperse phase premix may comprise avehicle containing cyclosporin and the surfactant. The vehicle may be ahydrophobic liquid, for example a liquid lipid, or it may be or comprisea material, for example a surfactant, for forming self-assemblystructures. In particular, a disperse phase premix may comprisecyclosporin A, the first surfactant, an oil and other oil solublecomponents for example an optional solvent. The premixes may contain oneor more surfactants suitable for the phase they are to form, aspreviously mentioned, for example the aqueous premix may comprise asecond surfactant.

The aqueous premix comprises, or usually consists of, a solution inwater of water-soluble constituents, namely the hydrogel-forming polymerand water-soluble excipient(s). The aqueous premix may include aplasticiser for the hydrogel-forming polymer, as described elsewhere inthis specification. The aqueous premix may include a second surfactant,e.g. to increase polymer viscosity and improve emulsification andthereby help prevent precipitation of active agent during processing.SDS is an example of such a surfactant. In any event, the constituentsof the aqueous premix may be agitated for a period sufficient todissolve/melt the components, for example, from 1 hour to 12 hours toform the completed aqueous premix.

The disperse phase pre-mix may comprise the first surfactant andcyclosporin as a dispersion or preferably a solution in a vehicle (forexample an oil phase) as described above, for example in a liquidcomprising an oil or in a liquid comprising component(s) ofself-assembly structures. For example an oil phase pre-mix may thereforebe a liquid lipid, for example a medium chain triglyceride (MCT)formulation, the medium chain triglyceride(s) being one or moretriglycerides of at least one fatty acid selected from C6-012 fattyacids, and cyclosporin A and the surfactant comprising or being a mediumor long chain fatty acid mono- or di-glyceride. Suitably an oil phasepre-mix is stirred at ambient temperature to form a solution of thecyclosporin in the oil and surfactant. In some embodiments, thecomponents of the oil phase premix are mixed (or otherwise agitated) fora period of, for example, 10 minutes to 3 hours to form the premix.

The two premixes may be combined and agitated, for example for a periodof a few seconds to an hour, for example from 30 seconds to 1 hour,suitably 5 mins to an hour, to form a dispersion of the disperse phasein an aqueous hydrogel-forming polymer to form the liquid composition ofthe invention. The dispersion may then be further processed to form thecomposition or a core. The two premixes may be combined into thedispersion by agitation in a mixing vessel; they may additionally oralternatively be combined in a continuous flow mixer.

The basic method for making a composition or core comprising cyclosporinand hydrogel-forming polymer matrix, therefore, is to mix a liquid form(preferably a solution) of the hydrogel-forming polymer (or mixture ofpolymers) with the cyclosporin, the surfactant (to avoid any ambiguitythe first surfactant) and the oil phase (and any other disperse phasecomponents) to form a dispersion in the polymer, which later in theprocess forms a hydrogel. The method normally comprises mixing togetheran aqueous polymer phase premix and a disperse phase premix. Takingaccount of the final composition required (as described elsewhereherein), the disperse phase pre-mix and the liquid hydrogel-formingpolymer (i.e. the solution or suspension of hydrogel-forming polymer,the aqueous phase) may be mixed in a weight ratio of from 1:1 to 1:10,particularly 1:4 to 1:9, e.g. 1:5 to 1:7. In general, only gentlestirring of the components is required using a magnetic or mechanicalsystem, e.g. overhead stirrer, as would be familiar to a person skilledin the art to achieve a dispersion of the disperse phase in the aqueousphase to form a colloid (which may be in the form of for example anemulsion or micro emulsion in which the aqueous hydrogel is thecontinuous phase). Continuous stirring is preferred. Mixing may also beachieved using an in-line mixing system. Any appropriate laboratorystirring apparatus or industrial scale mixer may be utilized for thispurpose for example the Magnetic Stirrer (manufactured by Stuart) orOverhead Stirrer (by KNF or Fisher). It is preferred to set up theequipment in such a way as to minimise evaporation of contents such as,for example, water. In one embodiment of the process of the invention,it is preferred to utilise a closed system for stirring in order toachieve this aim. In-line mixing may be particularly suitable for closedsystem processing. Suitably mixing of the two components takes place ata temperature of 50 to 70° C., or 55-75° C., e.g. 60-70° C.

The mixing of the two phases results in a colloid wherein the aqueoushydrogel-forming polymer is an aqueous continuous phase and thecomponent(s) not soluble in the aqueous phase are a disperse phase. Thecolloid may have the form of an emulsion or microemulsion.

In embodiments where the disperse phase is or comprises a secondsurfactant, the amount of the second surfactant may be selected suchthat, upon combination of the disperse phase premix with the aqueouspre-mix, the second surfactant concentration in the combined mixtureexceeds the CMC for the second surfactant used such that micelles areformed in the aqueous phase comprising the hydrogel-forming polymer.Depending on the concentration of second surfactant used, self-assemblystructures other than micelles may also form. The CMC for a particularsurfactant may be determined using well known methods, for example asdescribed in Surfactants and Polymers in Aqueous Solutions SecondEdition, Chapter 2, Holmberg et al. In embodiments mixing of the aqueousphase and a disperse phase which is or comprises a surfactant may resultin the formation of a clear liquid, for example a microemulsion, inwhich the aqueous phase comprising the hydrogel-forming polymer is thecontinuous phase. Microemulsions are a thermodynamically stabledispersion of self-assembly structures in the aqueous phase, the size ofthe self-assembly structures being sufficiently small to give atransparent appearance. The size of the self-assembly structures presentas the disperse phase resulting from the mixing of the aqueous andsurfactant phases may be from about 0.5 nm to 200 nm, for example about1 nm to 50 nm, or about 5 nm to 25 nm. The size of the self-assemblystructures formed and other characteristics such as the opticalisotropicity of the formulation (for example a microemulsion) may bedetermined using well known techniques such as dynamic light scattering.

Where the polymer matrix substantially consists of gelatin with theaddition of sorbitol, the aqueous phase of polymer matrix is prepared byadding the appropriate quantities of sorbitol (and surfactant ifdesired) to water, heating to approximately 50 to 75° C., for example60-75° C. until in solution and then adding gelatin, although theprecise order and timing of addition is not critical. A typical “gelatinsolution” comprises 8 to 35%, (for example 15-25%, preferably 17-18%)gelatin; 65%-85% (preferably 77-82%) of water plus from 1-5% (preferably1.5 to 3%) sorbitol. When present, a second surfactant (e.g. anionicsurfactant) in the aqueous phase premix may be present in an amount of0.1 to 5% (preferably 0.5 to 4%) wherein all parts are by weight of theaqueous phase.

Optionally the processing temperature required for a standard gelatincan be reduced to a desirable target temperature e.g. 37° C. by use oflower melting-point gelatin (or gelatin derivatives or mixtures ofgelatins with melting point reducers) or other polymer matrix materialsuch as, for example, sodium alginate. If gelatin droplets are beingformed by machine extrusion and immediately cooled, e.g. in a coolingbath, additional appropriate inlet tubing can be used to introduce anoil phase containing cyclosporin A at ambient temperature into thehotter fluid gelatin solution (and the mixture can be immediatelyhomogenized) very shortly before ejection from a beading nozzle or otherdropletting process such that the duration of exposure of thecyclosporin A to the higher temperature gelatin is limited so reducingthe degree of any heat-dependent degradation of the active ingredient.This process may use any appropriate device such as, for example, ahomogenizer, e.g. a screw homogenizer, in conjunction with anextrusion-type apparatus as described for example in WO 2008/132707(Sigmoid Pharma) the entirety of which is incorporated herein byreference. Alternatively, the aqueous- and oil-based solutions can bepumped at appropriate rates and passed through a static mixer to form anemulsion prior to dropping.

The colloid is formed by combining of the disperse phase premix with theliquid aqueous phase with stirring as described above. The resultantcolloidal dispersion then has the formulation of a solidified coredescribed above but with liquid water still present in the coreformulation.

Optionally the cyclosporin may be added after mixing the aqueous phaseand other components of a disperse phase of the type comprising a liquidlipid in addition to the cyclosporin, however, it is preferred that thecyclosporin is added together with the other components of the dispersephase as a premix.

The resulting colloid is then poured or introduced into a mould or othervessel or poured onto sheets or between sheets or delivered dropwise (orextruded) into another fluid such that the polymer matrix-containingaqueous phase, on solidification, takes the form of the mould, vessel,sheet or droplet/bead intended. It is preferred to progress tomould-forming e.g. beading without delay.

Solidification (gelling) can occur in a variety of ways depending on thepolymer of the matrix, for example by changing the temperature aroundthe mould, vessel, sheet, droplet/bead etc or by applying asolidification fluid or hardening solution so that the moulded shape isgelled or solidified. In certain embodiments both temperature change andapplication of a solidifying fluid or hardening solution are employedtogether or simultaneously. For example, when using a dropping methodsolidification can occur by dropping into cooling oil, air or acombination thereof.

In the preferred embodiment in which the composition or core takes theform of beads, the beads may be formed for example by dropping thecolloid dropwise into a fluid which effects solidification. Where theviscosity of the composition to be beaded reaches a certain point, dropformation becomes more difficult and specialised apparatus is thenpreferred.

By use of the term “dry”, it is not sought to imply that a drying stepis necessary to produce the dry core (although this is not excluded)rather that the solid or solidified aqueous external phase issubstantially free of water or free of available water. Solidificationof the aqueous phase (external phase) may have arisen through variousmeans including chemically (e.g. by cross-linking) or physically (e.g.by cooling or heating). In this respect, the term “aqueous phase” isnevertheless employed in this document to denote the external(continuous) phase of the core even though water, in certainembodiments, is largely absent from (or trapped within the cross-linkedmatrix of) the core. The external phase of the core is howeverwater-soluble and dissolves in aqueous media.

In the case where solidification can be achieved by raising or reducingtemperature, the temperature of the solidification fluid can be adaptedto achieve solidification of the core at a desired rate. For example,when gelatin is used as the hydrogel-forming polymer, the solidificationfluid is at a lower temperature than the temperature of the emulsionthus causing solidification, i.e. gelling, of the polymer matrix. Inthis case, the solidification fluid is termed a cooling fluid.

In the case where solidification can be achieved chemically, e.g. byinduction of cross-linking on exposure to a component of thesolidification fluid, the concentration of such component in thesolidification fluid and/or its temperature (or other characteristic orcontent) can be adjusted to achieve the desired rate and degree ofsolidification. For example, if alginate is chosen as the polymermatrix, one component of the solidification fluid may be acalcium-containing entity (such as, for example, calcium chloride) ableto induce cross-linking of the alginate and consequent solidification.Alternatively, the same or similar calcium-containing entity may beincluded (e.g. disperse) in the aqueous phase of the fluid emulsionprior to beading and triggered to induce cross-linking e.g. by applyinga higher or lower pH to a solidification fluid into which droplets ofemulsion fall dropwise or are introduced. Such electrostaticcross-linking can be varied as to the resulting characteristics of thebead by control of calcium ion availability (concentration) and otherphysical conditions (notably temperature). The solidification fluid maybe a gas (for example air) or a liquid or both. For example, whengelatin is used as the hydrogel-forming polymer matrix, thesolidification fluid can be initially gaseous (e.g. droplets passingthrough cooling air) and then subsequently liquid (e.g. droplets passinginto a cooling liquid). The reverse sequence may also be applied whilegaseous or liquid cooling fluids alone may also be used. Alternatively,the fluid may be spray-cooled in which the emulsion is sprayed into acooling gas to effect solidification.

In the case of gelatin or other water-soluble polymer (or polymermixture) destined to form an immobilization matrix, it is preferred thatthe solidification fluid be a non-aqueous liquid (such as, for example,medium chain triglycerides, mineral oil or similar preferably with lowHLB to ensure minimal wetting) which can conveniently be placed in abath (cooling bath) to receive the droplets of the colloid as theysolidify to form the beads of the core. Use of a non-aqueous liquidallows greater flexibility in choice of the temperature at which coolingis conducted.

Where a liquid cooling bath is employed, it is generally maintained atless than 20° C., preferably maintained in the range 5-15° C., morepreferably 8-12° C. when standard gelatin is used as thehydrogel-forming polymer. If a triglyceride is chosen as the coolingfluid in the cooling bath, a preferred example is Miglyol 840 fromSasol.

If alginate is selected as the polymer matrix, a typical method ofmaking beads involves dropwise addition of a 3% sodium alginate solutionin which oil droplets are disperse as described above into a 4° C.crosslinking bath containing 0.1 M calcium chloride to produce calciumalginate (this method can be referred to as “diffusion setting” becausethe calcium is believed to diffuse into the beads to effectcross-linking or setting). Using a syringe pump, or Inotech machine,droplets can be generated or extruded (egg at 5 mL/h if a pump is used)through a sterile needle or other nozzle (described elsewhere herein)which can be vibrating as discussed elsewhere herein. Airflow of between15 and 20 L/min through 4.5 mm tubing can be applied downwards over theneedle to reduce droplet size if desired. Newly formed beads can then bestirred in the calcium chloride bath for up to an hour. If carrageenanis used as the polymer matrix both salt and reduction in temperaturee.g. by dropping into cooling oil may be used to obtain solidification.

An alternative approach when using alginate is internal gelation inwhich the calcium ions are disperse in the aqueous phase prior to theiractivation in order to cause gelation of hydrocolloid particles. Forexample, this can be achieved by the addition of an inactive form of theion that will cause crosslinking of the alginate, which is thenactivated by a change in e.g. pH after sufficient dispersion of the ionis complete (see Glicksman, 1983a; Hoefler, 2004 which are bothincorporated herein by reference). This approach is particularly usefulwhere rapid gelation is desired and/or where the diffusion approach maylead to loss of API by diffusion thereof into the crosslinking bath.

Where another ionotropic polymer is used than alginate, suitableanalogous processes may be used to those described herein in relation toalginate.

Following shape-forming, moulding or beading, the resultant shapes orforms may be washed then dried if appropriate. In the case of beadssolidified in a solidification fluid, an optional final step in themethod of production described above therefore comprises removal of thesolidified beads from the solidification fluid. This may be achievede.g. by collection in a mesh basket through which the solidificationfluid (e.g. medium chain triglycerides) is drained and the beadsretained and is preferably conducted without delay e.g. as soon as thebeads have formed or within 5, 10, 15, 20, 25 or 30 minutes of theirformation. Excess solidification fluid may then be removed using acentrifuge (or other apparatus or machine adapted to remove excessfluid) followed by drying of the beads to remove water or free waterand/or removal of some or all of any additional solvent e.g. ethanol orisopropyl alcohol used to dissolve or facilitate dissolution of theactive principle in preceding steps optionally followed by washing (e.g.using ethyl acetate) and a subsequent “drying” step to remove excesssolvent (e.g. ethyl acetate). Isopropyl alcohol is an example of asolvent which is preferably removed later in processing to reduceresidues in the oil or aqueous phase. Drying can be achieved by anysuitable process known in the art such as use of a drum drier (e.g.Freund Drum dryer which may be part of the Spherex equipment train ifused) with warm air at between 15° C. and 25° C., preferably around 20°C. leading to evaporation or entrainment of the water by the air.Alternatively, drying may be carried out using of a fluid bed drier(e.g. Glatt GPCG 1.1) with warm air between 40° C. and 60° C.; or usinga vibrational fluid bed drier. Use of gelatin as the polymer matrix(e.g. as principal constituent of the aqueous immobilisation phase) inmost cases requires a drying step and for beads this is preferablyachieved by drying in air as above described. Beads can be dried usingother techniques, as understood by the person skilled in the art, forexample vibrating fluid bed, tray drying or vacuum drying. The resultantformulation (the formulation of the invention) is essentially dry asdescribed in more detail above.

In general, the beads may be generated by the application of surfacetension between the liquid dispersion (the mixture of the aqueous andsurfactant phases) and an appropriate solidification fluid such as, forexample, gas or liquid in order to create the spherical or substantiallyspherical shape of the ultimate beads.

Alternatively, the beads may be produced through ejection or extrusionof the liquid dispersion (the aqueous phase pre-mix and the dispersephase premix mixture) through an orifice or nozzle with a certaindiameter and optionally subject to vibration (using selected vibrationalfrequencies) and/or gravitational flow. Examples of machines which maybe used are encapsulation prilling, drop pelletising, spray cooling orspray congealing machines for example the Freund Spherex, ITAS/Lambo,Globex, Inotech, GEA Niro, Droppo, Buchi, Gelpell, Brace processingequipment processing equipment. Operation of the Spherex machinemanufactured by Freund as may be desired to manufacture beads accordingto the present invention is described in U.S. Pat. No. 5,882,680(Freund), the entire contents of which are incorporated herein byreference. It is preferred to select a vibrational frequency in theregion of 2-200 Hz, suitably 10-50 Hz, although the ultimate choice (andseparately the amplitude of vibration selected) depends on the viscosityof the dispersion to be beaded. If the polymer matrix is chosen tosolidify at lower temperature, it may be appropriate to maintain thelines to the orifice/nozzle at a certain temperature to maintain thefluidity of the solution. Suitably the colloid is ejected through asingle-orifice nozzle, e.g. having a diameter of from 0.1 mm to 5 mm(for example 0.5-5 mm), to form drops which are then caused or allowedto fall into a cooling oil or other hardening medium and allowed toharden to form seeds, after which the seeds are recovered from thecooling oil and dried.

It will be appreciated, therefore, that the invention includes a processfor manufacturing a composition of the invention or a core comprisingcyclosporin, a first surfactant being or comprising a medium chain orlong chain fatty acid mono- or di-glyceride or a combination thereof,wherein the surfactant does not comprise or is not a polyethyleneglycolether or ester, and an oil phase in a hydrogel forming polymer matrix,which process comprises: forming an aqueous premix which comprises waterand water soluble/dispersible materials (including therefore ahydrogel-forming polymer) and a disperse phase premix (e.g. an oil phasepremix) which comprises the oil phase, the cyclosporin and the firstsurfactant optionally other excipients (e.g. oil(s) and oilsoluble/dispersible materials), and combining the two premixes to form acolloid (disperse phase) within an aqueous phase comprising thehydrogel-forming polymer. The colloid may then be formed into a shapedunit, for example a bead to provide the core comprising the activeingredient. More particularly the manufacture of a composition or coreas defined above may comprise:

(i) forming an aqueous phase pre-mix comprising a solution in water ofwater-soluble constituents (e.g. of a hydrogel-forming polymer, anywater-soluble excipient(s), as described elsewhere herein);(ii) forming a disperse phase pre-mix typically comprising a dispersionor preferably a solution of cyclospori, in a liquid lipid, and the firstsurfactant, optionally together with other disperse phase constituents(e.g. surfactant, solvents etc as described elsewhere herein);(iii) mixing the aqueous phase pre-mix (i) and the disperse phasepre-mix (ii) to form a colloid;(iv) ejecting the colloid through a nozzle to form droplets;(v) causing or allowing the a hydrogel-forming polymer to gel orsolidify to form a water soluble polymer matrix; and(vi) drying the solid.

The manufacture of a liquid composition of the invention may comprise:

(i) forming an aqueous phase pre-mix comprising a solution in water ofwater-soluble constituents (e.g. of a hydrogel-forming polymer, anywater-soluble excipient(s), as described elsewhere herein);

(ii) forming a disperse phase pre-mix typically comprising a dispersionor preferably a solution of cyclospori, in a liquid lipid, and the firstsurfactant, optionally together with other disperse phase constituents(e.g. surfactant, solvents etc as described elsewhere herein); and

(iii) mixing the aqueous phase pre-mix (i) and the disperse phasepre-mix (ii) to form a colloid.

Some manufacturing processes comprise steps (A) to (D) below or,alternatively, a manufacturing process may comprise a single one or anycombination of steps (A) to (D).

(A) Exemplary Preparation of Aqueous Phase:

Aqueous phase components are added to water, e.g. purified water, underagitation e.g. sonication or stirring. The temperature is graduallyincreased, for example to 60-70° C. and in particular 65° C., to achievecomplete dissolution of the solids. The aqueous phase components includea hydrogel-forming polymer, e.g. gelatin or agar and optionally one ormore other excipients, for example D-sorbitol (a plasticiser) andsurfactant (for example SDS). Possible aqueous phase components aredescribed elsewhere herein.

The gelatin may be Type A gelatin. In some less preferredimplementations, the gelatin is Type B. The gelatin may have a Bloomstrength of 125-300, optionally of 200-300, for example of 225-300, andin particular 275. The components of the aqueous phase may be agitatedfor a period of, for example, from 1 hour to 12 hours to completepreparation of the aqueous phase (aqueous premix).

(B) Exemplary Preparation of Disperse Phase:

Cyclosporin is mixed with the first surfactant, an oil and otherdisperse phase components (for example a co-solvent) under agitatione.g. sonication or stirring, suitably at ambient temperature to disperseor preferably dissolve the active ingredient.

(C) Exemplary Mixing of the Two Phases

The aqueous phase and the disperse phase are mixed. The two phases maybe mixed in a desired weight; for example, the weight ratio of dispersephase to aqueous phase may be from 1:1 to 1:10, e.g. from 1:4 to 1:9 andoptionally from 1:5 to 1:8 such as about 1:5 or about 1:7. The resultingcolloid is agitated, e.g. sonicated or stirred, at a temperature of60-70° C. and in particular 65° C., to achieve a homogeneous dispersion,then the homogenous dispersion is formed into beads. In particular, thehomogenous dispersion is ejected through a single orifice nozzle to formdroplets which fall into a cooling medium. The nozzle is suitablyvibrated to facilitate droplet formation. The nozzle may be vibrated ata frequency of 2-200 Hz and optionally 15-50 Hz.

The cooling medium may for example be air or an oil; the oil is suitablyphysiologically acceptable as, for example, in the case of medium chaintriglycerides e.g. Miglyol 810N. The cooling medium may be at a coolingtemperature often of less than 15° C., for example of less than 10° C.but above 0° C. In some embodiments the cooling temperature is 8-10° C.The nozzle size (diameter) is typically from 0.5 to 7.5 mm, e.g. from0.5 to 5 mm and optionally from 0.5 to 4 mm. In some embodiments, thenozzle diameter is from 1 to 5 mm for example from 2 to 5 mm, andoptionally from 3 to 4 mm, and in particular may be 3.4 mm. The nozzlediameter may be from 1 to 2 mm.

The flow rate through a 3.4 mm nozzle or through a 1.5 mm nozzle is 5 to35 g/min and optionally 10 to 20 g/min and for nozzles of differentsizes may be adjusted suitably for the nozzle area.

(D) Exemplary Processing of Beads

Cooled beads are recovered, for example they may be recovered fromcooling oil after a residence time of 15-60 minutes, for example afterapproximately 30 minutes. Beads recovered from a cooling liquid (e.g.oil) may be centrifuged to eliminate excess cooling liquid, and thendried. Suitably, drying is carried out at room temperature, for examplefrom 15-40° C. and optionally from 20-35° C. The drying may be performedin a drum drier, for example for a period from 6 to 24 hours, e.g. ofabout 12 hours in the case of beads dried at room temperature. The driedbeads may be washed, suitably with a volatile non-aqueous liquid atleast partially miscible with water, e.g. they may be washed with ethylacetate. The washed beads may be dried at room temperature, for examplefrom 15-25° C. and optionally from 20-25° C. The drying may be performedin a drum drier, for example for a period from 6 to 48 hours, e.g. ofabout 24 hours in the case of beads dried at room temperature. Dryingmay be achieved by any suitable means, for example using a drum dryer,suitably under vacuum; or by simply passing warm air through the batchof beads, or by fluidising the beads in a suitable equipment with warmair, for example if a fluid bed dryer. Following drying, the beads arepassed through a 1 to 10 mm, optionally 2 to 5 mm to remove oversizedbeads and then through a sieve with a pore size of 0.5 to 9 mmoptionally 1 to 4 mm to remove undersized beads.

It can be appreciated that it is possible to recycle the beads that arerejected by the sieving process.

As a further aspect of the invention there is provided a formulationobtainable by (having the characteristic of) any of the processesdescribed herein. It is to be understood that the processes describedherein may therefore be used to provide any of the specific coresdescribed in embodiments herein by dispersing the appropriate componentswhich form the disperse phase of the core in the appropriate componentswhich form the aqueous continuous matrix phase of the core.

The preceding paragraphs describe the formation of uncoated compositionsor cores. The composition may comprise a coating. Cores may be coated.The composition or the core may be coated with a subcoat and/or coatedwith a second coating (also referred to as a modified release coating orouter coat). Suitable sub coats and modified release coatings (secondcoating or outer coat) are any of those described herein and any of thefirst coating (for the subcoat) or the second coating (for the modifiedrelease coating). The coating(s) may be applied using well knownmethods, for example spray coating as described below to give thedesired sub coat and modified release coating weight gains.

With regard to one of the methods described above (ejection of emulsionthrough an optionally vibrating nozzle) with two concentric orifices(centre and outer), the outer fluid may form a coating (outside thebead) as described herein. The Spherex machine manufactured by Freund(see U.S. Pat. No. 5,882,680 to Freund) is preferably used (the entirecontents of this patent is incorporated herein by reference). Othersimilar ejection or extrusion apparatus may also be used, for examplethe ejection apparatus described hereinbefore.

Use of the Spherex machine achieves very high monodispersity. Forexample, in a typical 100 g, batch 97 g of beads were between 1.4 to 2mm diameter or between 1 and 2 mm. Desired size ranges can be achievedby methods known in the art for rejecting/screening different sizedparticles. For example, it is possible to reject/screen out thelarger/smaller beads by passing a batch first through e.g. a 2 mm meshand subsequently through a 1.4 mm mesh.

The 1.4 to 2 mm diameter range is a good size if it is desired to spraycoat the beads (if smaller, the spray of the coating machine may bypassthe bead; if too large, the beads may be harder to fluidise, which isnecessary to achieve consistent coating).

Coating Process

The coating process can be carried out by any suitable means such as,for example, by use of a coating machine which applies a solution of apolymer coat (as described above in particular) to the formulation.Polymers for coating are either provided by the manufacturer inready-made solutions for direct use or can be made up before usefollowing manufacturers' instructions.

Coating is suitably carried out using a fluid bed coating system such asa Wurster column to apply the coating(s) to the composition or the core.Appropriate coating machines are known to persons skilled in the art andinclude, for example, a perforated pan or fluidized-based system forexample the GLATT, Vector (e.g. CF 360 EX), ACCELACOTA, Diosna, O'Haraand/or HICOATER processing equipment. To be mentioned is the MFL/01Fluid Bed Coater (Freund) used in the “Bottom Spray” configuration.

Typical coating conditions are as follows:

Process Parameter Values Fluidising airflow (m3/h) 20-60 (preferably30-60) Inlet air temperature (° C.) 20-65 Exhaust air temperature (° C.)20-42 Product temperature (° C.) 20-45 (preferably 40 to 42) Atomizingair pressure (bar) Up to 1.4 e.g. 0.8-1.2 Spray rate (g/min) 2-10 and3-25 RPM

Suitably the coating is applied as a solution or dispersion of thepolymers (and other components) of the coating. Generally the coatingsare applied as an aqueous, solution or dispersion, although othersolvent systems may be used if required. The coating dispersion isapplied to the composition or the core as a spray in the fluid bedcoater to give the required coating weight gain. Generally the coatingprocess is carried out at a temperature which maintains the cores at atemperature of from 35 to 45° C., preferably 40 to 42° C.

After applying the coating, the composition may be dried, for example bydrying at 40 to 45° C.

The invention further provides a product having the characteristics of acomposition obtained as described herein, a product defined in terms ofits characteristics being defined by the characteristics of thecomposition to the exclusion of the method by which it was made.

As mentioned herein the processes described may be used to provide anyof the composition described in the various embodiments herein. By wayof example there is provided a composition of the invention comprising acore and a first coating comprising a water-soluble cellulose ether or awater soluble derivative of a cellulose ether and/or a second coatingcomprising a delayed release polymer wherein the core comprises ahydrogel-forming polymer matrix comprising gelatin, cyclosporin A,medium chain mono-di- and/or tri-glycerides, a first surfactant being orcomprising a medium chain or long chain fatty acid mono- or di-glycerideor a combination thereof that does not comprise or is not apolyethyleneglycol ether or ester, a co-solvent and optionally a secondsurfactant, the core having the characteristics of a core obtained bythe process comprising steps (i) to (vi) described above for forming thecore, wherein the aqueous phase pre-mix in step (i) of the processcomprises gelatin and optionally a second surfactant (suitably ananionic surfactant), and the oil phase pre-mix in step (ii) of theprocess comprises medium chain mono-di- or tri-glycerides, hydrophobicactive ingredient, surfactant (suitably a non-ionic surfactant) andcosolvent; and the wherein the core is optionally coated with a firstcoating comprising a water-soluble cellulose ether or a water solublederivative of a cellulose ether and/or a second coating comprising adelayed release polymer; wherein the coatings are any of those describedherein. Accordingly, the process may produce a composition as describedabove comprising a first coating and/or a second coating. The processmay additionally produce a composition comprising a first coating and asecond coating being outside the first coating.

In the cores described herein to which the following characteristics areapplicable, e.g. in the immediately preceding paragraph, the followingcharacteristics may be present:

gelatin may be present in an amount of 300 to 700 mg/g;

the medium chain mono-, di- or tri-glycerides (for examplecaprylic/capric triglyceride) may be present in an amount of 20 to 200mg/g;

co-solvent (for example 2-(ethoxyethoxy)ethanol) may be present in anamount of 150 to 250 mg/g;

non-ionic surfactant (for example sorbitan-based surfactants, PEG-fattyacids, or glyceryl fatty acids or poloxamers or particularly apolyethoxylated castor oil for example Kolliphor EL) may be present inan amount of 80 to 200 mg/g;

anionic surfactant (for example, alkyl sulphates, carboxylates orphospholipids (particularly SDS)) may be present in an amount of 15 to50 mg/g; and

cyclosporin A, may be present in an amount of from 60 to 180 mg/g,suitably 60 to 150 mg/g, 90 to 150 mg/g, or 80 to 100 mg/g, for example81 to 98 mg/g;

wherein all weights are based upon the dry weight of the core beforecoating.

The composition may comprise or the core may be coated with a firstcoating (sub-coating) which is or comprises a water-soluble compoundselected from cellulose ethers and their derivatives, particularlyhydroxypropylmethyl cellulose; the first coating being present in anamount corresponding to a weight gain due to the first coating in arange selected from: (i) from 8% to 12%, for example about 10%; or (ii)from 4% to 6%, for example about 5% by weight based upon the weight ofthe core prior to applying the first coating. The first coating may havea modified release coating (or second coating) applied to it.

Preferably, any modified release coating (second coating), especially inthe embodiments of the immediately preceding paragraphs, is or comprisesa pH independent modified release coating, more especially the secondcoating may be a modified release coating comprising ethyl cellulose (egSurelease) still more particularly a modified release coating comprisingethyl cellulose and a water-soluble polysaccharide, pectin (e.g. aSurelease-pectin coating as described herein); and wherein the modifiedrelease coating is present in an amount corresponding to a weight gainof the formulation due to the second coating selected from (a) from 10%to 12%, for example about 11% or about 11.5%; or (b) from 16% to 18%,for example about 17% by weight based upon the weight of the formulationprior to applying the second coating.

In addition the process to form a composition of the invention maycomprise the steps of mixing a first population and a second population,wherein

the first population has a coating that is or comprises a water-solublecellulose ether but having no outer coating, e.g. as described herein;and

the second population has a first coating that is or comprises awater-soluble cellulose ether and a second coating that is or comprisesa delayed release coating, for example as described herein e.g. acoating that is or comprises a delayed release polymer.

Applications

The composition of the invention may advantageously be used for oraldelivery pharmaceutically active ingredients by virtue of the enhanceddissolution profiles achieved.

The compositions of the invention include modified release compositionswhich comprise cyclosporin A and a modified release coating, for examplecomprising a pH independent polymer, to target cyclosporin release tothe lower intestine. Such compositions result in low systemic exposureto cyclosporin A, whilst providing high levels of cyclosporin A in thelower GI tract, particularly in the colon. Such compositions release thecyclosporin A in an active form for example as a solution, whichprovides enhanced absorption of cyclosporin A in the local tissue of thelower GI tract. When the composition is used in the form of minibeads,the minibeads are advantageously dispersed along large sections of theGI tract following oral administration and are therefore expectedprovide a more uniform exposure to cyclosporin to large sections of forexample the colon.

Accordingly the modified release compositions according to the inventioncomprising cyclosporin for local treatment of the lower GI tract areexpected to be useful in the treatment or prevention of a condition ofthe GIT. In particular the composition of the invention may comprisecyclosporin A and/or another immunosuppressant and be useful in theprevention or treatment of inflammatory conditions affecting the lowerGI tract, particularly conditions affecting the colon.

The composition of the invention is administered orally. The doserequired will vary depending upon the specific condition being treatedand the stage of the condition. In the case of compositions containingcyclosporin A, the composition will generally be administered to providea dose of cyclosporin A of from 0.1 to 100 mg, for example a dose of 1to 500 mg or particularly a dose of 25 to 250 mg cyclosporin A. Thecomposition is suitably administered as a single daily dose.

In one aspect of the invention there is provided a composition of theinvention for use in the treatment or prophylaxis of an inflammatorybowel disease, Crohn's disease, ulcerative colitis, graft-versus-hostdisease, gastrointestinal graft-versus-host disease, myasthenia gravis,irritable bowel syndrome (e.g. with constipation, diarrhea and/or painsymptoms), celiac disease, stomach ulcers, diverticulitis, pouchitis,proctitis, mucositis, chemotherapy-associated enteritis,radiation-associated enteritis, short bowel disease, or chronicdiarrhea, gastroenteritis, duodenitis, jejunitis, peptic ulcer,Curling's ulcer, appendicitis, colitis, diverticulosis, endometriosis,colorectal carcinoma, adenocarcinoma, inflammatory disorders such asdiversion colitis, ischemic colitis, infectious colitis, chemicalcolitis, microscopic colitis (including collagenous colitis andlymphocytic colitis), atypical colitis, pseudomembraneous colitis,fulminant colitis, autistic enterocolitis, interdeminate colitis,jejunoiletis, ileitis, ileocolitis or granulomatous colitis, theprevention of rejection following bone marrow transplantation,psoriasis, atopic dermatitis, rheumatoid arthritis, nephrotic syndromeprimary sclerosing cholangitis, familial adenomatous polyposis, orperinanal Crohn's, including perianal fistulae.

In one embodiment the composition of the invention is for use in thetreatment of an inflammatory bowel disease. The main forms ofinflammatory bowel disease are Crohn's disease and ulcerative colitis.Accordingly the composition of the invention may be useful in thetreatment of both of these conditions.

The composition of the invention may be for use in the treatment orprevention of irritable bowel syndrome (e.g. with constipation, diarrheaand/or pain symptoms), celiac disease, stomach ulcers, diverticulitis,pouchitis, proctitis, mucositis, radiation-associated enteritis, shortbowel disease, or chronic diarrhea, gastroenteritis, duodenitis,jejunitis, peptic ulcer, Curling's ulcer, appendicitis, colitis,diverticulosis, endometriosis, colorectal carcinoma, adenocarcinoma,inflammatory disorders such as diversion colitis, ischemic colitis,infectious colitis, chemical colitis, microscopic colitis (includingcollagenous colitis and lymphocytic colitis), atypical colitis,pseudomembraneous colitis, fulminant colitis, autistic enterocolitis,interdeminate colitis, jejunoiletis, ileitis, ileocolitis, granulomatouscolitis, fibrosis, graft-versus-host disease, gastrointestinalgraft-versus-host disease, HIV prophylaxis and treatment (for exampleHIV enteropathy) or gastrointestinal enteropathies. The composition mayalso be for use in the treatment or prevention of Clostridium difficilecolitis.

Crohn's disease may affect the entire GI tract including the colon.However, ulcerative colitis is a condition which affects only the colonand the rectum. Accordingly, the release profile provided by thecolon-targeted, immunosuppressant-containing (e.g. cyclosporinA-containing), composition according to the invention is expected to beespecially beneficial in the treatment of ulcerative colitis.

The colon-targeted, composition of the invention primarily releasescyclosporin A, in the colon. However, cycosporin may also be releasedhigher in the GI tract and accordingly the composition may also providetherapeutic benefit in conditions which affect other parts of the lowerGI tract, for example Crohn's disease, irritable bowel syndrome (e.g.with constipation, diarrhea and/or pain symptoms), celiac disease,stomach ulcers, diverticulitis, collagenous colitis, pouchitis,proctitis, mucositis, radiation-associated enteritis, short boweldisease, chronic diarrhea, gastroenteritis, duodenitis, jejunitis,peptic ulcer, Curling's ulcer, appendicitis, diverticulosis,endometriosis, colorectal carcinoma, adenocarcinoma, inflammatorydisorders such as, jejunoiletis, ileitis, ileocolitis, celiac disease,fibrosis, graft-versus-host disease, gastrointestinal graft-versus-hostdisease, HIV prophylaxis and treatment (for example HIV enteropathy) orenteropathies.

Gastrointestinal Graft-Versus-Host-Disease (GI-GVHD) is alife-threatening condition and one of the most common causes for bonemarrow and stem cell transplant failure. In patients with GI-GVHD it isthe donor cells that begin to attack the patient's body—most frequentlythe gut, liver and skin. Patients with mild-to-moderate GI-GVHDtypically develop symptoms of anorexia, nausea, vomiting and diarrhoea.If left untreated, GI-GVHD can progress to ulcerations in the lining ofthe GI tract, and in its most severe form, can be fatal. Accordingly, inone embodiment the composition is for use in the treatment orprophylaxis of Gastrointestinal Graft-Versus-Host-Disease (GI-GVHD).

In a further embodiment there is provided a composition of the inventionfor use in the treatment of celiac disease.

In a further embodiment there is provided a composition of the inventionfor use in the treatment or prophylaxis of ulcerative colitis.

Also provided is a composition of the invention for use in the treatmentof neurodegenerative diseases (for example Parkinson's disease,Alzheimer's disease or vascular dementia) or paediatric diseases,including, but not limited to ulcerative colitis, Crohn's disease andGvHD.

Chronic inflammation of the GI tract may result in cellulartransformation and the onset of cancer through tumourigenesis.Cyclosporin has been shown to be effective in inhibiting cell growth ina number of colorectal cancer cell lines (Wereneck et al Cell Cycle11:21; 2012; 3997-4008). It has also been shown that cyclosporin may bean effective inhibitor of tumourigenesis (Kawahara et al, Cyclosporine Aand Tacrolimus Inhibit Urothelial Tumorigenesis; MolecularCarcinogenesis, 2015). Accordingly, cyclosporin may be beneficial inproviding a cytostatic anti-cancer effect thereby inhibiting the growthof a tumour. Cyclosporin may be beneficial in preventing or delaying theonset of colorectal cancer in patients with chronic inflammatoryconditions affecting the GI tract, particularly the colon, for examplethe inflammatory conditions described herein such as ulcerative colitisor Crohn's disease. As discussed above the compositions comprisingcyclosporin provide high levels of cyclosporin in a solubilised forminto the colon and may therefore by particularly beneficial in thetreatment of colorectal cancer.

A further aspect of the invention provides a composition comprisingcyclosporin as defined herein for use in the treatment of a canceraffecting the GI tract, particularly the lower GI tract and especiallythe colon. Accordingly the composition comprising cyclosporin may be foruse in the treatment of colorectal cancer. The composition comprisingcyclosporin may be for use in providing a cytostatic effect in a canceraffecting the GI tract, particularly a colorectal cancer.

Also provided is a composition comprising cyclosporin for use in thepreventing or delaying the onset of a cancer of the GI tract in apatient with chronic inflammatory condition affecting the GI tract,particularly the lower GI tract and especially the colon. Accordinglythe composition comprising cyclosporin may be for use in inhibitingtumourigenesis in the GI tract, particularly the colon.

The composition comprising cyclosporin may be used alone or togetherwith another anti-cancer agent, for example the composition comprisingcyclosporin may be used together with an antineoplastic agent to treator delay the onset of a cancer affecting the GI tract. The compositioncomprising cyclosporin may be administered to a subject as a fixed dosecombination with one or more additional anticancer agents. Thecomposition comprising cyclosporin may be administered separately,sequentially or substantially simultaneously with another anticanceragent.

Anti-cancer agents which may be suitable for use with the compositioncomprising cyclosporin include, but are not limited to one or moreagents selected from

(i) antiproliferative/antineoplastic drugs and combinations thereof,such as alkylating agents (for example cis-platin, oxaliplatin,carboplatin, cyclophosphamide, nitrogen mustard, uracil mustard,bendamustin, melphalan, chlorambucil, chlormethine, busulphan,temozolamide, nitrosoureas, ifosamide, melphalan, pipobroman,triethylene-melamine, triethylenethiophoporamine, carmustine, lomustine,stroptozocin and dacarbazine); antimetabolites (for example gemcitabineand antifolates such as fluoropyrimidines like 5-fluorouracil andtegafur, raltitrexed, methotrexate, pemetrexed, cytosine arabinoside,floxuridine, cytarabine, 6-mercaptopurine, 6-thioguanine, fludarabinephosphate, pentostatine, and gemcitabine and hydroxyurea); antibiotics(for example anthracyclines like adriamycin, bleomycin, doxorubicin,daunomycin, epirubicin, idarubicin, mitomycin-C, dactinomycin andmithramycin); antimitotic agents (for example vinca alkaloids likevincristine, vinblastine, vindesine and vinorelbine and taxoids liketaxol and taxotere and polokinase inhibitors); proteasome inhibitors,for example carfilzomib and bortezomib; interferon therapy; andtopoisomerase inhibitors (for example epipodophyllotoxins like etoposideand teniposide, amsacrine, topotecan, irinotecan, mitoxantrone andcamptothecin); bleomcin, dactinomycin, daunorubicin, doxorubicin,epirubicin, idarubicin, ara-C, paclitaxel (Taxol™), nabpaclitaxel,docetaxel, mithramycin, deoxyco-formycin, mitomycin-C, L-asparaginase,interferons (especially IFN-alpha), etoposide, teniposide,DNA-demethylating agents, (for example, azacitidine or decitabine); andhistone de-acetylase (HDAC) inhibitors (for example vorinostat, MS-275,panobinostat, romidepsin, valproic acid, mocetinostat (MGCD0103) andpracinostat SB939);(ii) cytostatic agents such as antiestrogens (for example tamoxifen,fulvestrant, toremifene, raloxifene, droloxifene and iodoxyfene),antiandrogens (for example bicalutamide, flutamide, nilutamide andcyproterone acetate), LHRH antagonists or LHRH agonists (for examplegoserelin, leuprorelin and buserelin), progestogens (for examplemegestrol acetate), aromatase inhibitors (for example as anastrozole,letrozole, vorazole and exemestane) and inhibitors of 5α-reductase suchas finasteride; and navelbene, CPT-II, anastrazole, letrazole,capecitabine, reloxafme, cyclophosphamide, ifosamide, and droloxafine;(iii) anti-invasion agents, for example dasatinib and bosutinib(SKI-606), and metalloproteinase inhibitors, inhibitors of urokinaseplasminogen activator receptor function or antibodies to Heparanase;(iv) inhibitors of growth factor function: for example such inhibitorsinclude growth factor antibodies and growth factor receptor antibodies,for example the anti-erbB2 antibody trastuzumab [Herceptin™] theanti-EGFR antibody panitumumab, the anti-erbB1 antibody cetuximab,tyrosine kinase inhibitors, for example inhibitors of the epidermalgrowth factor family (for example EGFR family tyrosine kinase inhibitorssuch as gefitinib, erlotinib,6-acrylamido-N-(3-chloro-4-fluorophenyl)-7-(3-morpholinopropoxy)-quinazolin-4-amine(CI 1033), erbB2 tyrosine kinase inhibitors such as lapatinib) andantibodies to costimulatory molecules such as CTLA-4, 4-IBB and PD-I, orantibodies to cytokines (IL-10, TGF-beta); inhibitors of the hepatocytegrowth factor family; inhibitors of the insulin growth factor family;modulators of protein regulators of cell apoptosis (for example Bcl-2inhibitors); inhibitors of the platelet-derived growth factor familysuch as imatinib and/or nilotinib (AMN107); inhibitors ofserine/threonine kinases (for example Ras/Raf signalling inhibitors suchas farnesyl transferase inhibitors, for example sorafenib, tipifarniband lonafarnib), inhibitors of cell signalling through MEK and/or AKTkinases, c-kit inhibitors, abl kinase inhibitors, PI3 kinase inhibitors,Plt3 kinase inhibitors, CSF-1R kinase inhibitors, IGF receptor, kinaseinhibitors; aurora kinase inhibitors and cyclin dependent kinaseinhibitors such as CDK2 and/or CDK4 inhibitors; and CCR2, CCR4 or CCR6antagonists;(v) antiangiogenic agents such as those which inhibit the effects ofvascular endothelial growth factor, [for example the anti-vascularendothelial cell growth factor antibody bevacizumab (Avastin™)];thalidomide; lenalidomide; and for example, a VEGF receptor tyrosinekinase inhibitor such as vandetanib, vatalanib, sunitinib, axitinib andpazopanib;(vi) gene therapy approaches, including for example approaches toreplace aberrant genes such as aberrant p53 or aberrant BRCA1 or BRCA2;(vii) immunotherapy approaches, including for example antibody therapysuch as alemtuzumab, rituximab, ibritumomab tiuxetan (Zevalin®) andofatumumab; interferons such as interferon α; interleukins such as IL-2(aldesleukin); interleukin inhibitors for example IRAK4 inhibitors;cancer vaccines including prophylactic and treatment vaccines such asHPV vaccines, for example Gardasil, Cervarix, Oncophage and Sipuleucel-T(Provenge); gp100; dendritic cell-based vaccines (such as Ad.p53 DC);toll-like receptor modulators for example TLR-7 or TLR-9 agonists; PD-1,PD-L1, PD-L2 and CTL4-A modulators (for example Nivolumab), antibodiesand vaccines; other IDO inhibitors (such as indoximod); anti-PD-1monoclonal antibodies (such as MK-3475 and nivolumab); anti-PDL1monoclonal antibodies (such as MEDI-4736 and RG-7446); anti-PDL2monoclonal antibodies; and anti-CTLA-4 antibodies (such as ipilumumab;and(viii) cytotoxic agents for example fludaribine (fludara), cladribine,pentostatin (Nipent™);

The coating containing the water-soluble cellulose ether of the presentinvention may be useful in reducing the variability between releaseprofiles of different batches of minibeads.

A “batch” is a specific quantity of a drug or other material that isintended to have uniform character and quality, within specified limits,and is produced according to a single manufacturing order during thesame cycle of manufacture. A “lot” means a batch, or a specificidentified portion of a batch, having uniform character and qualitywithin specified limits; or, in the case of a drug product produced bycontinuous process, it is a specific identified amount produced in aunit of time or quantity in a manner that assures its having uniformcharacter and quality within specified limits. “Lot number”, “controlnumber”, or “batch number” means any distinctive combination of letters,numbers, or symbols, or any combination of them, from which the completehistory of the manufacture, processing, packing, holding, anddistribution of a batch or lot of drug product or other material can bedetermined.”

EXAMPLES Example 1a: Preparation of a Liquid Composition of theInvention

An aqueous phase was prepared by mixing sodium dodecyl sulphate (SDS)and D-sorbitol with purified water under constant stirring. Gelatin wasthen added to this solution and gentle heat was applied to approximately60-70° C. to achieve complete melting of gelatin. The composition of theaqueous phase is shown in Table 1 below.

TABLE 1 Component w/w % water 79.6 SDS 1.3 Sorbitol 2.0 Gelatin 17.1

An oil phase was prepared by mixing together 2-(2-ethoxyethoxy)ethanol(Transcutol HP), glyceryl monooleate/dioleate (Capmul GMO-50) andcapric/caprylic triglyceride (Miglyol 810) with stirring at roomtemperature to form a solution. Ciclosporin A was added and mixed untila clear solution was obtained. The composition of the oil phase is shownbelow in Table 2.

TABLE 2 Component w/w % Cyclosporin A 24.5 Miglyol 810 N 12.5 TranscutolHP 37.0 Capmul GMO-50 26

The oil phase was mixed with the heated aqueous phase in a ratio ofapproximately 1:5 (oil phase:aqueous phase). The resulting mixture wasstirred at 60-70° C., 250-350 rpm using a magnetic stirrer to achievehomogeneity.

Example 1b: Preparation of a Further Liquid Composition

Following the procedure of Example 1a a further liquid composition withglyceryl caprylate/caprate (Capmul MCM) as the surfactant in the oilphase in place of the glyceryl monooleate/dioleate (Capmul GMO-50) wasprepared. The aqueous phase of the composition is shown in Table 3 andthe oil phase of the composition is shown in Table 4. The oil phase wasmixed with the heated aqueous phase in a ratio of approximately 1:5 (oilphase:aqueous phase).

TABLE 3 Component w/w % water 79.6 SDS 1.3 Sorbitol 2.0 Gelatin 17.1

TABLE 4 Component w/w % Cyclosporin A 24.5 Miglyol 810 N 12.5 TranscutolHP 37.0 Capmul MCM 26

Example 1c: Preparation of a Further Liquid Composition

Following the procedure of Example 1a a further liquid composition withglycerol linoleate (Maisine 35-1) as the surfactant in the oil phase inplace of the glyceryl monooleate/dioleate (Capmul GMO-50) was prepared.The aqueous phase of the composition is shown in Table 5 and the oilphase of the composition is shown in Table 6. The oil phase was mixedwith the heated aqueous phase in a ratio of approximately 1:5 (oilphase:aqueous phase).

TABLE 5 Component w/w % water 79.6 SDS 1.3 Sorbitol 2.0 Gelatin 17.1

TABLE 6 Component w/w % Cyclosporin A 24.5 Miglyol 810 N 12.5 TranscutolHP 37.0 Maisine 35-1 26

Example 2: Preparation of a Minibead

A minibead as described herein may be a composition of the invention.Alternatively the minibead may be a core. The minibead was generallyprepared by forming a minibead according to the following procedure

The composition or core in the form of seamless minibeads were preparedusing Spherex process as follows.

An aqueous phase and oil phase mixture was prepared following theprocedure described in Example 1a.

The mixture was then fed (via temperature controlled tubing) through avibrating nozzle, with a single nozzle outlet with a diameter of 3 mm.Seamless minibeads were formed as the solution flowed through thevibrating nozzle into a cooling chamber of constantly flowing mediumchain triglyceride (Miglyol 810) cooling oil at a temperature of 10° C.

The minibeads were removed from the cooling oil and placed in acentrifuge to remove the excess oil. Following centrifugation, a firstdrying step was initiated with a set refrigerator temperature of 10° C.and the heater temperature of 20° C. The dryer was rotated at 15 RPM.When the beads were observed to be freely rotating in the drying drum,they were considered to be dry.

The minibeads were washed with ethyl acetate and then dried for afurther 24 h under the same drying conditions as those mentioned abovein the first drying step. The dried minibeads were then sieved to removeoversize and undersize beads resulting in cores 1 mm-2 mm in diameter.This procedure provided cores with the composition shown in Table 7, thevalues being the weight percent of the total weight for each component.

TABLE 7 Component w/w % Cyclosporin A 12.1 Miglyol 810 N 6.2 TranscutolHP 18.3 Capmul GMO-50 12.9 SDS 3.2 Sorbitol 4.9 Gelatin 42.4

Example 3: Preparation of a Minibead with a First Coating (Sub-Coat)

A coated minibead can be produced by coating a minibead produced inExample 2 with a dispersion of Opadry White 20A28380 (supplied byColorcon). The minibeads were loaded into a fluid bed coater (Wurstercolumn) and coated with Opadry White 20A28380 (supplied by ColorconLimited) as a dispersion. The processing parameters, such as inlet airtemperature and inlet air volume, were adjusted to keep the minibeadtemperature between 40° C. and 42° C. until the required coating weightgain was reached. The resulting subcoated minibeads were dried for 5minutes at 40° C. in the coater.

Composition of the Coated Minibead

A minibead with the composition shown in Table 8 below was produced bythe above procedure. A minibead with an Opadry weight gain of 7.5%relative to the weight of the core is shown in Table 8. Table 9 showsthe composition of a minibead coated with an Opadry weight gain of 5%relative to the weight of the core. Table 10 shows the composition of aminibead coated with an Opadry weight gain of 10% relative to the weightof the core.

TABLE 8 Component w/w % Cyclosporin A 11.3 Miglyol 810 N 5.8 TranscutolHP 17.0 Capmul GMO-50 12.0 SDS 2.9 Sorbitol 4.6 Gelatin 39.4 Opadry 7.0

TABLE 9 Component w/w % Cyclosporin A 11.5 Miglyol 810 N 5.9 TranscutolHP 17.4 Capmul GMO-50 12.3 SDS 3.1 Sorbitol 4.7 Gelatin 40.3 Opadry 4.8

TABLE 10 Component w/w % Cyclosporin A 11.0 Miglyol 810 N 5.6 TranscutolHP 16.7 Capmul GMO-50 11.7 SDS 2.9 Sorbitol 4.5 Gelatin 38.5 Opadry 9.1

Example 4a: Preparation of a Minibead with a Second Coating ofEthylcellulose

A minibead coated with Opadry, the first coating (also referred to as asubcoat), was produced following the procedure in Example 3. Theminibead produced by the procedure of Example 3 was then further coatedwith a second coating (also referred to as an overcoat) of Surelease®(an ethylcellulose dispersion).

The Surelease® overcoat was applied by the following procedure.Surelease® was slowly added to a stainless steel vessel and mixed toprovide the required coating suspension of Surelease® for the overcoat.The resulting coating suspension was then applied onto the surface ofthe sub-coated minibeads using an analogous coating method to thatdescribed for the Opadry coating in Example 3 until the desired weightgain of Surelease® was reached. The over-coated minibeads were thendried in the coater for an hour at 40-45° C.

The minibead was coated with a 9.5% weight gain of Surelease®.

Minibeads with no Opadry coating may be produced by coating a minibeaddescribed in Example 2 with Surelease® as described above.

The minibead with a first and second coating has the composition shownin Table 11.

TABLE 11 Component w/w % Cyclosporin A 10.3 Miglyol 810 N 5.3 TranscutolHP 15.5 Capmul GMO-50 10.9 SDS 2.7 Sorbitol 4.2 Gelatin 36.0 Opadry 6.4Surelease 8.7

Similarly, the composition of minibeads coated with 5% Surelease and7.5% Opadry are shown in Table 12 and the composition of minibeadscoated with 20% Surelease and 7.5% Opadry are shown in Table 13.

TABLE 12 Component w/w % Cyclosporin A 10.7 Miglyol 810 N 5.5 TranscutolHP 16.2 Capmul GMO-50 11.4 SDS 2.9 Sorbitol 4.4 Gelatin 37.5 Opadry 6.6Surelease 4.8

TABLE 13 Component w/w % Cyclosporin A 10.7 Miglyol 810 N 5.5 TranscutolHP 16.2 Capmul GMO-50 11.4 SDS 2.9 Sorbitol 4.4 Gelatin 37.5 Opadry 6.6Surelease 4.8

Example 4b: Preparation of a Minibead with a Second Coating ofEthylcellulose/Pectin

A minibead coated with Opadry, the first coating (also referred to as asubcoat), was produced following the procedure in Example 3. Theminibead produced by the procedure of Example 3 was then further coatedwith a second coating (also referred to as an overcoat) of a mixture ofSurelease® (an ethylcellulose dispersion) and Pectin.

The Surelease®/pectin overcoat was applied by an analogous method to theSurelease coating of Example 4a. Pectin was added to purified water in astainless steel vessel and mixed to obtain a solution. Surelease® wasslowly added to the vessel whilst maintaining mixing to provide therequired Pectin concentration in the Surelease® for the overcoat. Theresulting coating suspension was then applied onto the surface of thesub-coated minibeads using an analogous coating method to that describedfor the Opadry coating in Example 3 until the desired weight gain ofSurelease®/Pectin was reached. The over-coated minibeads were then driedin the coater for an hour at 40-45° C.

The minibead was coated with a 9.5% weight gain of Surelease®/Pectin.

The minibead with a first and second coating has the composition shownin Table 14.

TABLE 14 Component w/w % Cyclosporin A 10.3 Miglyol 810 N 5.3 TranscutolHP 15.5 Capmul GMO-50 10.9 SDS 2.7 Sorbitol 4.2 Gelatin 36.0 Opadry 6.4Surelease 8.5 Pectin 0.2

Example 5: Crystallisation Experiments Comparative Example 1

Three liquid compositions comprising Cremophore EL (also known asKremophore EL), a polyethoxylated castor oil surfactant with an HLBvalue of greater than 10 were prepared. The three liquid compositionsare different sublots from the same batch. Each of the liquidcompositions has an oil phase comprising: cyclosporin A 26.3%,Transcutol HP 40%, Cremophor EL 22.5%, and Miglyol 810 11.2% (% amountsare of the oil phase); and an aqueous phase comprising: gelatin 17.1%,Sorbitol 2.0%, SDS 1.4%, and water 79.5% (% amounts are of the aqueousphase. The liquid composition was prepared by mixing the oil phase andaqueous phases in an oil phase to aqueous phase ratio of 1:7.

Example 5a

Three liquid compositions of Example 1a comprising Capmul GMO-50,replacing the Cremophore EL were prepared. The three liquid compositionsare 3 sublots of the same batch. Capmul GMO-50 has an HLB value of 3.

Example 5b

A liquid composition of Example 1 b comprising Capmul MCM was preparedCapmul MCM has an HLB value of 6-7.

Example 5c

A liquid composition of Example 1c comprising Maisine 35-1 was prepared.Maisine 35-1 has an HLB value of

The crystallisation rate of the liquid compositions of ComparativeExample 1, Example 5a, Example 5b and Example 5c were tested todetermine the length of time for cyclosporin crystallisation to occur.Each of the liquid compositions was stirred at 250-350 rpm to form anemulsion. Samples of the three emulsions were taken at 30 minuteintervals and viewed under a microscope at 50× or 100× magnification.The time when crystals appeared in the sample is shown in Table 15.

TABLE 15 Crystallization Beads Example Surfactant HLB time (h) formation5a Capmul GMO-50 3 2 Yes 5b Capmul MCM 5-6 >7 No 5c Maisine 35-1 4 3 YesComparative Cremophor EL 14  0.5 Yes Ex. 1

FIGS. 1A-1I show images of the three liquid compositions of ComparativeExample 1 comprising Cremophore EL (also known as Kremophore EL), apolyethoxylated castor oil surfactant with an HLB value of greater than10.

FIGS. 2A-21 show images of three liquid compositions of Example 5acomprising Capmul GMO-50 which is replacing the Cremophore EL. CapmulGMO-50 has an HLB value of 3. The three compositions comprising CapmulGMO-50 are

FIGS. 3A-3J show images of a liquid composition of Example 5b,comprising Capmul MCM.

FIGS. 4A-4J show images of a liquid composition of Example 5c comprisingMaisine 35-1. As is evident from the images the liquid compositions ofExample 5a, 5b and 5c had a much longer period before crystals appeared.The Capmul GMO-50 and Capmul MCM compositions were essentially free ofcrystal formation throughout the test period. The Capmul GMO-50compositions were essentially free of crystal up to 240 min, whereas theCremophore EL compositions had noticeable crystal formation after 120min. The Capmul MCM compositions were crystal free for 7 hours. TheMaisine 35-1 composition had crystal formation at 3 hours.

Example 6: In-Vitro Dissolution Profile of Minibeads of Example 2

The in-vitro dissolution profiles of a sample of the minibeads producedin Example 2 were measured using the following dissolution test. Thedissolution testing was carried out in accordance with USP <711>Dissolution using Apparatus II (paddle apparatus) operated with a paddlespeed of 75 rpm and with the dissolution medium at a temperature of 37°C.±0.5° C. The dissolution medium was deionised water. The sample ofminibeads were placed in the dissolution medium and at the start of thetest (t=0). The dissolution medium was sampled at regular intervals. Theobtained dissolution data is shown in the Table 16 and Table 17 below.The dissolution profile for two batches of the minibead of Example 2 isshown in FIG. 5.

TABLE 16 Timepoint ESN-740 ESN-740 ESN-740 ESN-740 ESN-740 ESN-740 %(Hrs) Pot 1 Pot 2 Pot 3 Pot 4 Pot 5 Pot 6 RSD 0.08 18.7 21.7 18.3 19.716.5 15.4 12.2 0.17 49.3 51.6 49.1 49.9 45.2 36.6 11.7 0.25 67.0 67.072.6 67.4 64.2 54.0 9.5 0.5 76.4 77.1 79.0 76.2 89.5 63.7 10.7 1 81.582.6 84.0 85.5 77.7 77.2 4.1 2 92.1 90.5 87.2 95.4 82.4 87.9 5.0 4 101.595.7 85.5 97.4 82.6 93.2 7.8 6 99.5 97.1 91.2 98.7 96.2 94.8 3.1 12 98.794.8 94.8 90.2 91.9 95.5 3.1 24 100.5 99.1 52.3 85.3 93.1 88.0 20.5

TABLE 17 Timepoint ESN-760 ESN-760 ESN-760 ESN-760 ESN-760 ESN-760 %(Hrs) Pot 1 Pot 2 Pot 3 Pot 4 Pot 5 Pot 6 RSD 0.08 16.1 15.5 18.2 20.217.5 6.4 30.8 0.17 47.6 44.2 52.3 51.4 44.2 24.2 23.4 0.25 65.6 61.768.7 69.6 57.3 34.6 21.9 0.5 75.7 71.2 75.2 75.9 68.0 45.7 17.0 1 78.978.0 79.0 84.8 76.4 61.6 10.2 2 94.6 90.7 97.9 97.9 90.7 68.9 12.1 4102.7 97.5 98.2 101.0 97.9 82.3 7.6 6 102.2 99.1 97.5 102.1 101.0 84.07.1 12 104.5 99.1 100.0 94.9 99.5 94.9 3.6 24 102.0 100.2 101.0 100.0102.7 101.2 1.0

Comparative Example 2

Minibeads corresponding to those of Example 2 were prepared withCremophore EL as the surfactant in place of Capmul GMO-50. Theseminibeads had the composition shown in Table 18. These minibeads weresubmitted to the same dissolution test in deionised water. Thedissolution profile of 5 batches of these minibeads is shown in FIG. 6.

TABLE 18 Component w/w % Cyclosporin A 10.8 Miglyol 810 N 4.6 TranscutolHP 16.4 Cremophor EL 9.2 SDS 4.0 Sorbitol 5.8 Gelatin 49.2

Example 6a

As a further comparison with Comparative Example 2, minibeads wereprepared using almost identical quantities of each excipient to those ofComparative Example 2, except that the 9.2 w/w % Cremophor was replacedwith 9.3% Capmul GMO-50 to give the minibead composition shown in Table19. The minibeads of Table 19 were prepared in an analogous manner tothose of Example 2. The w/w % in Tables 18 and 19 refer to the dryweight of the composition.

TABLE 19 Component w/w % Cyclosporin A 10.9 Miglyol 810 N 4.6 TranscutolHP 16.6 Capmul GMO-50 9.3 SDS 4.0 Sorbitol 5.7 Gelatin 49.0 CyclosporinA 10.8

These minibeads were submitted to the same dissolution test in deionisedwater. The dissolution profile of 3 batches of these minibeads is shownin FIG. 7.

It is apparent from the dissolution profiles shown in FIGS. 5, 6 and 7that compositions comprising Capmul GMO-50 (FIGS. 5 and 7) have superiordissolution profiles compared to compositions comprising Cremophor EL(FIG. 6). The dissolution profile of the Capmul GMO-50 compositions havea higher maximum release and this maximum release of cyclosporin isgenerally maintained in solution for longer than that observes withComparative Example 2, which contained Cremophor. FIG. 6 shows that the% release of cyclosporin from the Cremophor EL compositions is lower andthat the cyclosporin concentration reduces over time from a maximumcompared to the compositions containing Capmul GMO-50.

Example 7: In-Vitro Dissolution Profile of Minibeads of Example 4a

Minibeads corresponding to those of Example 4a, specifically minibeadswith the composition shown in Table 11 (7.5% Opadry subcoat, firstcoating and 9.5% Surelease overcoat, second coating) were prepared,having Capmul GMO-50 as the surfactant. These minibeads were submittedto a 2 stage dissolution test.

In the first stage of the test the dissolution medium was 750 ml of 0.1N HCl simulating the pH of the gastric environment. At the start of thetest (t=0) the sample was placed in the dissolution medium. After 2hours an aliquot of the medium is taken for subsequent analysis andimmediately (suitably within 5 minutes) the second stage of thedissolution test is initiated. In the second stage 250 ml of 0.2Mtribasic sodium phosphate containing 2% sodium dodecyl sulphate (SDS) isadded to the dissolution medium and the pH adjusted to 6.8±0.05 using 2NNaOH or 2N HCl as required.

Samples of the dissolution medium were taken at the following timepoints during the second stage of the test: 4 hours; 6 hours; 12 hours;and 24 hours from the start of the test (i.e. from t=0 at the start ofthe first stage).

The sample taken at the end of the first stage (2 hours) and the samplesfrom the second stage were analysed for cyclosporin A using ReversePhase HPLC with UV detection at 210 nm.

The amount of dissolved cyclosporin A in the dissolution medium isexpressed as a percentage based upon the original cyclosporin content inthe test formulation (the % released). The percentage release is givenin Table 21 and the dissolution profile of minibeads with thecomposition shown in Table 11 is shown in FIG. 8.

TABLE 21 Timepoint % Drug release (hours) Batch 1 Batch 2 Batch 3 Batch4 0 0 0 0 0 2 0 0 0 0 4 15 15 15 17 6 39 38 36 40 12 70 69 64 71 24 9999 95 100

Example 8: Droplet Size—Dynamic Light Scattering

The size of the droplets was measured using dynamic light scattering.Coated minibeads of Example 4a, Table 11 (minibeads coated with 7.5% wtgain Opadry and 9.5% wt gain Surelease) (0.5 g) comprising Capmul GMO-50as the first surfactant were added to a beaker containing 50 g ofdeionised water. The beaker contents were mixed at 250 rpm throughoutthe study. Samples of the beaker contents were taken at 0, 1, 2, 3, 4,5, 6 and 24 hours.

Samples of the beaker contents were filtered using 0.65 μm pore sizefilters (Merck Millipore Ultrafree-CL Centrifugal Filter). The particlesize and zeta potential of each sample was measured and analysed using aMalvern Nano-Zetasiser. The resulting data is shown in FIG. 9.

The coated minibeads tested in this example exhibit very stable dropletrelease for up to 4 hours with droplet size ranging from 120 to 240 nmin water media. As time passes and further droplets are released fromthe coated minibeads after 4 hours the droplet size range is broader.Without wishing to be bound by theory, it is possible that as timepasses and the dissolution media becomes more saturated with releaseddroplets a re-equilibration between the droplets already present in themedia and the ones freshly released may occur. The variability indroplet size after 4 hours is potentially caused by thisre-equilibration process rather than a representation of the size of thedroplets being released from the minibeads.

Example 9: In-Vivo Study in Healthy Male Volunteers

In the study described below “CyCol®” is a reference to the minibeads ofExample 4a, and described in Table 11 (comprising a core with CapmulGMO-50 as the surfactant, a 7.5% weight gain Opadry® first coating onthe core, and a 9.5% weight gain of a Surelease® second coating). Theminibeads were loaded into HPMC capsules to provide a unit dose of 37.5mg cyclosporin A per capsule.

LIST OF ABBREVIATIONS AND DEFINITION OF TERMS Abbreviation DefinitionAUC_(% extrap) Residual area/Percentage of AUC_(0-inf) extrapolatedAUC_(0-inf) Area under the concentration-time curve from time zero toinfinity (extrapolated) AUC_(0-t) Area under the concentration-timecurve from time zero to the last non-zero concentration AUC_(last) Areaunder the concentration-time curve from time zero to last quantifiableconcentration C_(av) Average steady state concentration CYP CytochromeP-450 k_(el) Elimination rate constant LLQ Lower limit of quantificationLR Linearity ratio SD Standard deviation t_(1/2) Terminal half-life UCUlcerative colitis

Study Objectives Primary Objectives:

-   -   To characterise whole blood pharmacokinetics of CyCol® following        single and multiple oral doses, and compare to a single        Sandimmun® IV administration pharmacokinetic profile in healthy        male subjects.    -   To evaluate the colonic mucosa (epithelial, mucosal and        sub-mucosal tissue) concentrations of cyclosporin and its        metabolites following multiple oral doses of CyCol® and compare        to concentrations following a single Sandimmun® IV        administration.

Secondary Objectives:

-   -   To obtain safety and tolerability information following multiple        oral doses of CyCol® at the selected doses in healthy male        subjects.

Exploratory Objectives:

-   -   To evaluate the amount of unchanged cyclosporin and its        metabolites excreted in the faeces after administration of        multiple doses of CyCol® and compare to amounts following a        single Sandimmun® IV administration.

Investigational Plan Overall Study Design and Plan—Description

This was a Phase I, single centre, multi-stage open study designed toevaluate the safety, tolerability, pharmacokinetics and relative colonicmucosal concentrations of cyclosporin capsules (CyCol®) compared to IVcyclosporin in healthy male volunteers. This study also investigated theamount of unchanged cyclosporin recovered from faecal samples andrelative concentrations of its metabolites AM9, AM4N and AM1. Theconcentrations of cyclosporin metabolites were also examined in faecaland colonic mucosa samples.

In Stage 1 of the study, a total of 24 eligible subjects received eitherSandimmun® IV over 24 hours (two consecutive 12 hour infusions) at adose of 2 mg/kg (2 mg/kg/day), a once daily oral dose of CyCol® 75 mgfor 7 days or a twice daily (BID) oral dose of CyCol® 75 mg for 7 days.Subjects who received CyCol® 75 mg BID only received a single dose onDay 7 (morning dose).

At the end of Stage 1 the data were reviewed and based on the evidenceavailable, the protocol allowed for further investigations to beconducted with alternative CyCol® doses and dose frequencies insubsequent stages. Following this review, 8 subjects were recruited toStage 2 of the study and received a once daily oral dose of CyCol® 37.5mg for 7 days. A further 8 subjects were recruited to Stage 3 andreceived a BID oral dose of CyCol® 150 mg for 7 days.

Details of the dosing sequence are presented in FIG. 10.

Study Design, Including Choice of Control Groups

A multi stage design was used for this study to reduce the number ofdose levels investigated and the number of subjects exposed tocyclosporin and study procedures. The dosing regimens chosen for Stages2 and 3 were chosen following review of data (safety and tolerability,systemic exposure and colonic mucosa tissue concentrations) observed atother doses.

The study was open-label because of the different mode of administrationof the comparator product (IV versus oral for investigational medicinalproduct), and the objective endpoints of the study (cyclosporinconcentrations). The single dose pharmacokinetic profile was examinedover a 24 hour period, based on previous experience that demonstratedthis duration adequately characterised the concentration-time profile.

The study recruited healthy male volunteers. Females were not includedin this study as the Food and Drug Administration pregnancy category forcyclosporin is C.

The comparator chosen for this study was Sandimmun®. The currenttreatment regimen for UC can often involve the use of several agentsadministered rectally, orally or intravenously depending on the severityof the disease. Cyclosporin is unlicensed for UC. However, according tothe 2013 National Institute for health and Clinical Excellenceguidelines for UC, it is recommended that treatment with IV cyclosporinshould be considered for subjects with acute severe colitis and notresponding to or unsuitable for first line therapy with corticosteroids.An IV dose of 2 to 4 mg/kg/day or an oral dose of 5 to 8 mg/kg/day isrecommended for severe ulcerative colitis treatment. The rationale forchoosing IV administration of cyclosporin over the oral formulation isbased on studies documenting variable absorption and the extensive firstpass metabolism following oral ingestion. In addition, an IV dose of 2to 4 mg/kg/day is known to be efficacious in the treatment of patientswith UC and to attain the same concentrations using a currentlyavailable oral dose form, approximately 3 times the IV dose would berequired. Furthermore, when cyclosporin is administered by IV infusionblood concentrations are constant and metabolites are present in lowerconcentrations compared to oral administration. CyCol® has beendeveloped to delay the release of active cyclosporin until it reachesthe colon, it thereby bypasses the site of absorption in the jejunum andthus limits the amount of metabolism (both pre-systemic metabolism inthe gastrointestinal tract and systemic metabolism by the hepaticsystem) by cytochrome P450 enzymes, including the CYP3A4 enzyme.Therefore, the concentration of metabolites following CyCol®administration is expected to be similar to IV cyclosporin. Furthermore,colon tissue concentrations of cyclosporin are 10 times higher inhealthy volunteers given cyclosporin parentally compared to those ofhealthy volunteers given the drug orally.

Treatments Treatments Administered

Subjects who met all the eligibility criteria for Stage 1 of theprotocol were assigned to one of the following dose groups on Day 1:

-   -   Sandimmun® 2 mg/kg IV infusion over 24 hours (2 mg/kg/day).    -   CyCol® 75 mg (2×37.5 mg CyCol® capsules) once daily orally for 7        days.    -   CyCol® 75 mg (2×37.5 mg CyCol® capsules) BID orally for 7 days        (only a single [morning] dose was administered on Day 7).

Following review of the Stage 1 data, the following CyCol® dosingregimen was explored in Stage 2:

-   -   CyCol® 37.5 mg (1×37.5 mg CyCol® capsule) once daily orally for        7 days.

Following review of the data from Stages 1 and 2, the following CyCol®dosing regimen was explored in Stage 3:

-   -   CyCol® 150 mg (4×37.5 mg CyCol® capsule) BID orally for 7 days        (only a single [morning] dose was administered on Day 7).

The dosing regimens chosen for Stages 2 and 3 were based upon safety andtolerability, systemic exposure and colonic tissue concentrationsobserved at other doses. To investigate higher doses the lower doses hadto have been well tolerated and had to have maximum systemic exposure250 ng/ml. Providing these criteria were met, doses could have beenescalated higher in search of a regimen that yielded colonic tissueconcentrations >300 ng/ml.

Identity of Investigational Products

Sandimmun® was provided in commercial packaging of 1 mL ampoulescontaining 50 mg of cyclosporin. The 1 mL ampoules of Sandimmun® werediluted with normal saline in accordance with the Summary of ProductCharacteristics (SmPC) and prepared suitably for the two consecutive 12hour infusions. Each infusion was prepared such that the total doseadministered was 2 mg/kg/day.

Methods of Assigning Subjects to Treatment Groups

In Stage 1, subjects were assigned sequentially to study treatment(Sandimmun® IV, CyCol® 75 mg once daily or CyCol® 75 mg BID).

All subjects recruited to Stage 2 received CyCol® 37.5 mg once dailyorally for 7 days and all subjects recruited to Stage 3 received CyCol®150 mg BID orally for 7 days.

Selection and Timing of Dose for each Subject

Sandimmun®

On the morning of Day 1, following an overnight fast, subjects startedtheir IV infusion via an IV catheter. Sandimmun® IV was administeredover 24 hours, as two consecutive 12 hour infusions, at a dose of 2mg/kg/day according to preparation and administration detailed in theSandimmun® SmPC.

CyCol®

On the morning of Day 1, following an overnight fast, subjects assignedto the CyCol® groups were administered the morning dose withapproximately 240 mL of water. A hand and mouth check was performed toensure ingestion of the study drug. Time of dosing was set to the timethe first capsule was administered.

On Day 2, prior to discharge from the study centre, subjects receivedthe morning dose with approximately 240 mL of water. For those subjectsassigned to the BID dosing regimen, the pharmacy dispensed the eveningdose to the subject with instructions to take the study drug asprescribed and within 10 to 12 hours following the morning dose.

Subjects returned to the study centre daily to receive the morning doseof CyCol® until Day 6 when the subjects were readmitted for an overnightstay. For those subjects assigned to the BID dosing regimen, at eachdaily visit to the study centre, the pharmacy dispensed the evening doseto subjects with instructions for the evening dose administration.

If the subject was expected to return to the study centre on Day 6 afterthe scheduled time of the Day 6 morning dose, the pharmacy could havedispensed the Day 6 morning dose to the subject upon discharge from thestudy centre on Day 5, with instructions for the morning doseadministration.

The last dose of CyCol® administration was on the morning of Day 7 forall subjects.

Blinding: This was an Open Label Study. PharmacokineticAssessments—Whole Blood

Blood samples for pharmacokinetic analysis were collected from allsubjects on Day 1 at 0 (pre dose), 2, 3, 4, 5, 6, 8, 10, 12, 16, 20 and24 hours post-dose (post start of infusion for the Sandimmun® IV group).

For subjects in the Sandimmun® IV group blood samples were alsocollected at 2, 4, 6 and 8 hours after completion of the infusion on Day2.

For subjects in the CyCol® groups trough pharmacokinetic samples wereobtained pre morning dose on Day 4. For subjects who received CyCol®once daily additional pharmacokinetic samples were obtained at 6, 12 and16 hours post morning dose on Day 6. For subjects who received CyCol®BID additional pharmacokinetic samples were obtained at 6 and 12 hourspost morning dose (prior to evening dose) on Day 6, and 4 hours postevening dose. On Day 7, all subjects in the CyCol® groups had bloodsamples for pharmacokinetics obtained at 0 (pre dose), 2, 4, 6 8, and 12hours post-dose. Actual sampling times were used for statisticalanalyses and so each time was recorded accurately.

Flexible Sigmoidoscopy and Colon Biopsies

All sigmoidoscopies were performed in an unprepared bowel (except forair and water), unless the subject had not defaecated within 24 hours ofsigmoidoscopy.

Subjects who received Sandimmun® IV were required to have thesigmoidoscopy performed within the last hour of their infusion (infusionhad to be on-going).

Subjects who received CyCol® were required to have the sigmoidoscopyperformed within 4 to 6 hours of the Day 7 morning/last dose.

The sigmoidoscopy was conducted by an appropriately trained endoscopistwho was familiar with the study protocol and obtaining biopsies. Perprotocol, whenever possible, the same endoscopist was to perform all thesigmoidoscopies. In this study two endoscopists performed thesigmoidoscopies.

For subjects who had not defaecated within 24 hours of thesigmoidoscopy, a rectal enema (Fleet saline, or similar) wasadministered 15 minutes before the scheduled biopsy time and the timeand volume of enema administration was recorded. The time and totalweight of the voided gut contents post enema were recorded. Arepresentative sample (approximately 5 g) was obtained from the voidedgut for processing as described herein.

The colon biopsies were collected (as described below) as soon aspossible and ideally within 10 minutes of the enema being voided.

Standard pinch biopsy forceps were used to obtain the colonic mucosabiopsies. Each biopsy was approximately 5 mm in size (and includedmucosa and submucosa layers). A total of 5 biopsies, approximately 1 cmapart were obtained from as close to the sigmoid colon as possible. Thetime of the biopsy collection and distance from the anal verge to theregion where biopsies were obtained were recorded in the CRF.

In the event that access to the sigmoid colon was limited, the 5biopsies could have been obtained from the rectum.

Each biopsy was rinsed with saline, blot dried and then transferred to apre-weighed collection tube. The tube was weighed to enabledetermination of the biopsy weight. The biopsy, without any furtherpreparation or processing was transferred to a cryovial and stored at−70° C. and transported to the bioanalytical laboratory on dry ice.

Concentrations of cyclosporin and its metabolites in the mucosal tissuewere assessed by a bioanalytical laboratory. Analysis of the tissuesamples was carried out as described below under “Colonic TissueAnalysis”

Faecal Sample Collection

Subjects were recommended to defaecate upon admission to the studycentre on Day 0. This sample was collected and one aliquot was collectedas a blank matrix.

From Day 0 through to completion of the study, subjects were requestedto collect their faeces. For subjects who received Sandimmun® IV,samples collected during the infusion were kept separate from thosecollected after completion of the infusion.

The time and date of each sample were recorded. Each sample wascollected and weighed. Faecal samples were homogenised as soon aspossible following collection.

Subjects who produced faecal samples as an outpatient were instructed tocollect the samples in the appropriate containers and store the samplesat room temperature until return to the study centre.

After homogenisation one aliquot of approximately 5 g was collected fromeach sample and frozen at −70° C. (with no additives).

The three intracolonic samples (approximately 500 mg to 1 g), ifavailable, were collected and stored in individual containers withoutany additives at −70° C.

For subjects who were administered a rectal enema, the time and totalweight of the voided gut contents post enema were recorded. Arepresentative sample (approximately 5 g) was obtained from the voidedgut and transferred to a container without any additives or additionalprocessing. All samples were stored at −70° C. and transported to abioanalytical laboratory on dry ice.

Drug Concentration Measurements

Blood samples were collected as described herein for determination ofcyclosporin concentrations. The amount of unchanged cyclosporin andrelative concentrations of its metabolites (AM9, AM4N and AM1) in faeceswere also assessed using faecal samples collected as described herein.

Primary Endpoints Whole Blood Pharmacokinetics

The following single dose pharmacokinetic parameters were derived afterdosing on Day 1 for both the Sandimmun® IV and CyCol® groups usingstandard non-compartmental methods:

-   -   C_(max): maximum observed concentration    -   T_(max): time of observed C_(max)    -   AUC_(0-t): area under the concentration-time curve from time        zero to the last non-zero concentration, determined using the        linear/log trapezoidal method    -   AUC_(0-inf): area under the concentration-time curve from time        zero to infinity (extrapolated) determined by        AUC_(0-t)+(C_(last)/k_(el)), where C_(last) is the predicted        concentration at the last quantifiable time point estimated from        the log-linear regression analysis.    -   t_(1/2): elimination half-life calculated as log_(e)(2)/k_(el)    -   k_(el): elimination rate constant (derived to calculate        AUC_(0-inf)), determined by a linear regression of the        log-linear concentration-time curve. Only those data points        judged to describe the terminal log-linear decline were used in        the regression    -   Residual area (% extrapolated; AUC_(% extrap)): calculated as        100*(1−AUC_(0-t)/AUC_(0-inf)) and derived to quantify the area        extrapolated to infinity    -   AUC_(0-t): area under the concentration-time curve over the        dosing interval (derived to calculate the accumulation ratio        discussed below).

The following steady state parameters were derived for the CyCol®groups:

-   -   C_(max), T_(max), AUC_(0-t) and average steady state        concentration (C_(av))

Additionally the accumulation ratio was derived to estimate theaccumulation over the dosing interval from single dose to steady state.

AC=AUC_(0-t)(steady state)/AUC_(0-t)(single dose)

The linearity ratio (LR) was derived to estimate the relative exposureper dose with steady state compared to a single dose.

LR=AUC_(0-t)(steady state)/AUC_(inf)(single dose)

For the Sandimmun® IV group simulations to steady state were conductedby the clinical pharmacokineticist to predict the concentration timeprofile and estimate C_(max), T_(max) and AUC_(0-T) for the group (i.e.,group values, not individual values).

To determine the steady state CyCol® parameters both the Day 6 and Day 7concentrations were used as summarised below.

Time Days used Days used post-dose for once daily for BID 0 7 7 2 7 7 47 6, 7 6 6, 7 6, 7 8 7 7 12 6, 7 6, 7 16 6 NA 24 Day 7 pre-dose NA

Actual sampling times were used in the derivation of parameters.

Cyclosporin concentrations were listed and summarised by group, day andnominal time post-dose.

Individual subject and median profiles of the concentration-time data(by day and by group) were plotted by dose group using actual andnominal times respectively. Median profiles were plotted on bothlinear-linear and log-linear.

Each pharmacokinetic parameter was summarised by group and by day. Forthe single dose, pharmacokinetic parameters, the Day 1 parameters weresummarised combining the once daily and BID groups using the same dose.

To assess the relationship between the pharmacokinetic parameters anddose, dose normalised AUC_(0-inf), AUC_(last) and C_(max) were plottedagainst dose for Day 1 and dose normalised C_(av) and C_(max) wereplotted against dose for Day 7 (using a logarithmic scale). These plotsincluded individual subject values and the geometric means for eachdose. The values were dose normalised (to a 1 mg dose) by dividing theindividual values and raw geometric means by dose.

Colonic Mucosal Tissue and Mucous Layer Pharmacokinetics

Concentrations of cyclosporin and its metabolites (AM9, AM4N and AM1) inboth the colonic mucosal tissue and mucous layer were determined. Forthe Sandimmun® IV group simulations were conducted by the clinicalpharmacokineticist to estimate the steady state concentrations.

Concentrations of cyclosporin and its metabolites (AM9, AM4N and AM1)were listed. Concentrations were also plotted against distance from theanal verge.

The concentration of cyclosporin in the colonic tissue was measured asfollows.

Colonic Tissue Analysis

Concentrations of cyclosporin and its metabolites (AM1, AM9 and AM4N) incolonic tissue was determined using the following protocol:

Principle

Liquid-liquid extraction with internal standardisation and HPLCseparation using a C18-column, followed by MS/MS detection.

Internal Standard—D12—Cyclosporin A Sample Matrix—Human Tissue

Calibration standards and quality control samples are prepared in 50%EtOH.

Solutions

The IS stock solution and respective dilutions are prepared by usingDMSO/MeOH (1/1). The internal standard (IS) working solution is preparedby dilution of the IS stock solution or one of its dilutions withDMSO/MeOH (1/1), and should have a concentration of ˜50 ng/mL

Storing of Samples and Solutions

Samples/solutions should be stored at −20° C. to −80° C.

Sample Handling and Sample Preparation for Analysis StepThawing/transfer procedure (step by step) 1 The following thawingprocedures are possible: Thawing at approximately 20 to 25° C. in awater bath for approx. 10 minutes Thawing air exposed at approximately20 to 25° C. for at least 30 minutes (depends on sample volume) 2 Ifapplicable: Vortexing for 30 seconds  3a Cal. Stds. & QCs: Transfer of1000 μL of each sample into a sample vial  3b Study sample: weight:approx.. 2-20 mg 4 Re-freezing of original samples between −20° C. and−80° C. Unless used for immediate preparation −> freezing of transferredsamples between −20° C. and −80° C.

Chromatographic and Auto-Sampler Parameters Parameter Scheduledrange/description Mobile phase 10 mM Ammonium acetate in water solvent AMobile phase ACN/THF (8/2) solvent B Mobile phase 10 mM Ammonium acetatein water solvent loading pump Chromatographic run 0.0-4.5 min lineargradient: 40% B → 52% B 4.5-6.0 min linear gradient: 52% B → 85% B6.0-6.01 min linear gradient: 85% B → 0% B 6.01-7.0 min isocratic: 0% BFlow 0.8 mL/min Injection volume 10 μL Pre-column/Column Luna C18, 4 × 2mm/ACE3AQ; 100 × 2.1 mm, 3 μm (ACT, UK) Column temperature 80° C.Cooling set point (T) 25° C.

Detection Parameter Scheduled range/description MS Ionisation mode ESIMS polarity Positive MS detection mode MRM Vaporizer temperature 600° C.Ionisation voltage 5.5 kV Gas 1 Pressure = 75 psi Gas 2 Pressure = 75psi Curtain gas pressure = 40 psi Lateral position 5 units ± 2 units(default) Vertical position 4 units ± 2 units (ESI default) Quadrupoleresolution low → low Transitions 1203.0 ± 0.3 → 99.9 ± 0.3 m/z:Cyclosporin A (CE: 125 eV, CXP: 16 V) 1215.0 ± 0.3 → 99.9 ± 0.3 m/z:D12-Cyclosporin A (CE: 125 eV, CXP: 16 V) 1219.0 ± 0.3 → 224.0 ± 0.3m/z: AM1 (CE: 65 eV, CXP: 15 V) 1219.0 ± 0.3 → 99.9 ± 0.3 m/z: AM9 (CE:125 eV, CXP: 16 V) 1189.0 ± 0.3 → 224.0 ± 0.3 m/z: AM4N (CE: 65 eV, CXP:15 V) DP (declustering potential) 130 V ± 20 V

Acceptance Criteria for Chromatograms Scheduled range/acceptanceParameter criteria/description AM1 Retention time for SST 4.2 min ± 0.5min AM4N Retention time for SST 5.4 min ± 0.5 min AM9 Retention time forSST 4.4 min ± 0.5 min

Calibration Standards and Quality Control Samples: Preparation of BlankSamples and Processed Matrix:

-   -   Preparation as described below, but taking DMSO/MeOH (1/1)        instead of IS working solution.

Step Preparation procedure (step by step) I [if not stored/available as1000 μL aliquots already −> see transfer above] II [if frozen −> thawingat 20° C. to 25° C. in a water bath for approx.. 5 min] 1 Addition of 25μL of internal standard working solution 2 Addition of 4 mL of DIPE 3Conversion Point: Extraction by shaking the test tubes vigorously forapprox. 5 minutes using a DVX-2500 Multi-tube Vortexer (1700 rpm; cycle:5 seconds run, 1 second pause time) 4 Centrifugation (phase separation)at 4000 rpm for 2 minutes 5 Storage at −75° C. for about 10 minutes 6Decanting of the organic, liquid phase into a centrifuge vial 7Evaporation of the organic phase using compressed air (Turbovap) atabout 40° C. for 14 minutes 8 Addition of 50 μL of 50% EtOH 9 Vortexingfor approx. 2 minutes using a DVX-2500 Multi-tube Vortexer (2500 rpm;cycle: 5 seconds run, 1 second pause time) 10 Centrifugation at 4000 rpmfor 1 minute

Carry-Over Samples:

-   -   Transfer of approx. 100 μL 50% EtOH into appropriate        auto-sampler vials        Matrix Samples (Human Tissue)—part A:

Step Preparation procedure (step by step) I [if not stored/available asapprox.. 2-20 mg aliquots already −> see transfer above] II [if frozen−> thawing at 20° C. to 25° C. in a water bath for approx.. 5 min]  1Addition of 500 μL 2% N-Acetyl-L-Cysteine in water  2 Vortexing forapprox. 10 min using a DVX-2500 Multi-tube Vortexer (1000 rpm)  3Centrifugation (phase separation) at 13000 rpm for 2 minutes usingbiofuge pico  4 Decanting of the liquid phase into a sample vial(volume: approx. 10 mL)  4a Caution: The remaining residue will beprepared separately (described in part B)  5 Addition of 500 μL EtOH tothe liquid phase  9 Addition of 25 μL of internal standard workingsolution 10 Addition of 4 mL of DIPE 11 Conversion Point: Extraction byshaking the test tubes vigorously for approx. 5 minutes using a DVX-2500Multi-tube Vortexer (1700 rpm; cycle: 5 seconds run, 1 second pausetime) 12 Centrifugation (phase separation) at 4000 rpm for 2 minutes 13Storage at −75° C. for about 10 minutes 14 Decanting of the organic,liquid phase into a centrifuge vial 15 Evaporation of the organic phaseusing compressed air (Turbovap) at about 40° C. for 14 minutes 16Addition of 50 μL of 50% EtOH 17 Vortexing for approx. 2 minutes using aDVX-2500 Multi-tube Vortexer (2500 rpm; cycle: 5 seconds run, 1 secondpause time) 18 Centrifugation at 4000 rpm for 1 minuteMatrix Samples (Human Tissue)—Part B (Samples are Taken from Part A,Step 4a):

Step Preparation procedure (step by step) 1 Addition of 500 μL 50% EtOHto the remaining residue 2 Addition of 25 μL of internal standardworking solution 3 Destroying of the tissue by using an ultrasonicprocessor (cycle: 0.5 s, max. amplitude) for 30 s 4 Decanting of theliquid phase into a sample vial (volume: approx. 10 mL) 5 Addition of500 μL 50% EtOH to the remaining residue to the remaining residue 6Vortexing for approx. 1 min using a DVX-2500 Multi-tube Vortexer (2500rpm; cycle: 5 seconds run, 1 second pause time) 7 Decanting of theliquid phase including all tissue into the same sample vial as used instep 4 8 Addition of 4 mL of Diisopropylether (DIPE) 9 Conversion Point:Extraction by shaking the test tubes vigorously for approx. 5 minutesusing a DVX-2500 Multi-tube Vortexer (1700 rpm; cycle: 5 seconds run, 1second pause time) 10 Centrifugation (phase separation) at 4000 rpm for2 minutes 11 Storage at −75° C. for about 10 minutes 12 Decanting of theorganic, liquid phase into a centrifuge vial 13 Evaporation of theorganic phase using compressed air (Turbovap) at about 40° C. for 14minutes 14 Addition of 50 μL of 50% EtOH 15 Vortexing for approx. 2minutes using a DVX-2500 Multi-tube Vortexer (2500 rpm; cycle: 5 secondsrun, 1 second pause time) 16 Centrifugation at 4000 rpm for 1 minute

Faceal Analysis

The concentration of unchanged cyclosporin and the relativeconcentrations of the metabolites (AM9, AM4N and AM1) were reported foreach intracolonic faecal sample and the faeces collected over theduration of the study.

If an enema was used prior to sigmoidoscopy then the appropriate faecalsample had the reported concentrations adjusted for the extra weight ofenema used (multiply by [total faecal weight/total-enema]).

Following collection of sigmoidoscopy biopsy samples, three intracolonicfaecal samples were taken from the region of the biopsy collection siteto test for cyclosporin concentrations.

Amounts of unchanged cyclosporin and the metabolites were plotted foreach subject against the collection time. The amount per hour wascalculated and plotted for the collection interval. Hence the plotconsists of stepped lines where the area under each step equates to theamount of drug measured for the collection interval.

Times since doses taken since the start of the collection interval werelisted.

The concentrations from the intracolonic faecal samples taken after thesigmoidoscopy were listed with the colonic mucosal tissue and mucouslayer concentrations.

Determination of Cyclosporin-A and its Metabolites, AM1, AM9 and AM4N,in Faecal Samples

The faecal samples were analysed by the RP-LC-MS/MS method using theprotocol below.

Method: Preparation of Solutions and Validation Samples ConcentratedSolutions and Dilutions

To spike calibration standards, quality control samples and othercontrol samples, concentrates and dilutions were prepared as shown inthe following table using the reference items and the internal standardwith purities as described above.

Solutions Used for Preparation Conc. [μg/mL] Name/date PreparationCyclosporin A AM1 AM4N AM9 K2718-1640 5.27 mg of Cyclosporin A weredissolved 500.00 — — — in 10.3819 mL of DMSO/MeOH (1/1) K2733-1634 2.57mg of AM1 were dissolved in 10 mL — 251.86 — — of DMSO/MeOH (1/1)K2731-1649 1.76 mg of AM4N were dissolved in 10 — — 149.60 — mL ofDMSO/MeOH (1/1) K2732-1648 5.03 mg of AM9 were dissolved in 10 mL — — —367.19 of DMSO/MeOH (1/1) V1-B-812 420 μL of K2718-1640, 417 μL ofK2733-  21.000  10.503  10.502  10.502 1634, 702 μL of K2731-1649, and286 μL of K2732-1648 were put together and filled up to 10 mL withDMSO/MeOH (1/1) K2711-1577 1.9 mg of D12-Cyclosporin A wereD12-Cyclosporin A dissolved in 10 mL of DMSO/MeOH (1/1) V1-IS-3-811 500μL of K2711-1577 were filled up to 9.2720 10 mL of DMSO/MeOH 1/1IS-WS-1-814- 865 μL of V1-IS-3-811 were mixed with 0.16013 Faeces 50 mLof DMSO/MeOH 1/1

Calibration Standards and Quality Control Samples

For analytical calibration purposes calibration standards and qualitycontrol samples were spiked with either a defined volume of aconcentrated solution described above or a higher concentratedcalibration standard into 50% Ethanol at eight concentrationlevels/three concentration levels respectively:

Preparation of Calibration Standards Volume of added Matrix solutionAdded volume Conc. [ng/mL] of Std [μL] solution Matrix [mL] CyclosporinA AM1 AM4N AM9 Std0B-812 — — 50% 4 — — — — Std1B-812 102.6 Std4B-812Ethanol 4 2.00 1.00 1.00 1.00 Std2B-812 65.2 Std5B-812 4 4.00 2.00 2.002.00 Std3B-812 48.6 Std8B-812 4 12.0 6.00 6.00 6.00 Std4B-812 348Std8B-812 4 80.0 40.0 40.0 40.0 Std5B-812 48.1 V1-B-812 4 250 125 125125 Std6B-812 97.6 V1-B-812 4 500 250 250 250 Std7B-812 148.2 V1-B-812 4750 375 375 375 Std8B-812 200 V1-B-812 4 1000 500 500 500 QC-A2-812187.4 QC-B2-812 50% 6 5.00 2.50 2.50 2.50 QC-B2-812 63.4 V1-B-812Ethanol 8 165 82.6 82.6 82.6 QC-C2-812 338 V1-B-812 8 851 426 426 426

Other Control Samples

Preparation of Other Control Samples (about 1 mg Blank Faeces added peraliquot) Volume of added Matrix solution Added volume Conc. [ng/mL] ofStd [μL] solution Matrix [mL] Cyclosporin A AM1 AM4N AM9 QC-A2-812 187.4QC-B2-812 50% 6 5.00 2.50 2.50 2.50 QC-B2-812 63.4 V1-B-812 Ethanol 8165 82.6 82.6 82.6 QC-C2-812 338 V1-B-812 8 851 426 426 426

Sample Preparation for Analysis (Processing) Calibration Standards andQuality Control Samples:

Preparation of blank samples and (if applicable) processed matrix:

-   -   Preparation as described below, but taking DMSO/MeOH (1/1)        instead of IS working solution.

Step Preparation procedure (step by step) 1 Addition of 0.4 mL of water2 Addition of 25 μL of internal standard working solution 3 Addition of4 mL of DIPE 4 Extraction by shaking the test tubes vigorously forapprox. 5 minutes using a DVX-2500 Multi-tube Vortexer (1700 rpm; cycle:5 seconds run, 1 second pause time) 5 Centrifugation (phase separation)at 4000 rpm for 2 minutes 6 Storage at −75° C. for about 10 minutes 7Decanting of the organic, liquid phase into a centrifuge vial 8Evaporation of the organic phase using compressed air (Turbovap) atabout 40° C. for 14 minutes 9 Addition of 750 μL of 50% EtOH 10Vortexing for approx. 2 minutes using a DVX-2500 Multi-tube Vortexer(2500 rpm; cycle: 5 seconds run, 1 second pause time) 11 Centrifugationat 4000 rpm for 1 minute 12 Transfer of an volume adequate to injectionpurposes into appropriate auto-sampler vials 13 Crimping the vials withappropriate vial caps

Matrix Samples (Human Faeces):

Step Preparation procedure (step by step) I [if not stored/available as100 mg aliquots already] II [if frozen −> thawing at 20° C. to 25° C. ina water bath for approx.. 5 min] 1 Fill up the volumetric flask (5 mL)with 50% EtOH 2 Vortexing for about 1 min using a vortex mixer 3 Wait(settle down) for 3 min 4 Transfer of 50 μL into a sample vial 5Addition of 950 μL 50% EtOH 6 Vortexing for approx. 30 s using aDVX-2500 Multi-tube Vortexer (2500 rpm) 7 Transfer of 50 μL into asample vial 8 Addition of 0.4 mL of water 9 Addition of 25 μL ofinternal standard working solution 10 Addition of 4 mL of DIPE 11Extraction by shaking the test tubes vigorously for approx. 5 minutesusing a DVX-2500 Multi-tube Vortexer (1700 rpm; cycle: 5 seconds run, 1second pause time) 12 Centrifugation (phase separation) at 4000 rpm for2 minutes 13 Storage at −75° C. for about 10 minutes 14 Decanting of theorganic, liquid phase into a centrifuge vial 15 Evaporation of theorganic phase using compressed air (Turbovap) at about 40° C. for 14minutes 16 Addition of 750 μL of 50% EtOH 17 Vortexing for approx. 2minutes using a DVX-2500 Multi-tube Vortexer (2500 rpm; cycle: 5 secondsrun, 1 second pause time) 18 Centrifugation at 4000 rpm for 1 minute 19Transfer of an volume adequate to injection purposes into appropriateauto-sampler vials 20 Crimping the vials with appropriate vial caps

Apparatus

Instruments and Materials Instrument/material Code Manufacturer Workstation API 6500 Mass Spectrometer 6500 Q-Trap AB SCIEX, USA/Canada

Software Data acquisition Analyst 1.6.2 (AB SCIEX, USA/Canada) Dataprocessing Analyst 1.6.2 (AB SCIEX, USA/Canada) Statistics andcalculations Analyst 1.6.2 (AB SCIEX, USA/Canada) Lotus 123 (Lotus Corp,USA)

Chromatographic Conditions and Detection Parameters

Chromatographic Conditions Parameter Scheduled range/description Mobilephase 10 mM Ammonium acetat in water solvent A Mobile phase ACN/THF(8/2) solvent B Chromatographic run 0.0-4.5 min linear gradient: 40% B →52% B 4.5-6.0 min linear gradient: 52% B → 85% B 6.0-6.01 min lineargradient: 85% B → 0% B 6.01-7.0 min isocratic: 0% B Flow 0.8 mL/minInjection volume 10 μL Injector flush DMSO/MeOH/Water (1/1/1)Pre-column/Column Luna C18, 4 × 2 mm/ACE3AQ; 100 × 2.1 mm, 3 μm (ACT,UK) Column temperature 80° C. Cooling set point (T) 25° C.

Detection Parameters Parameter Scheduled range/description MS Ionisationmode ESI MS polarity Positive MS detection mode MRM Vaporizertemperature 600° C. Ionisation voltage 5.5 kV Gas 1 Pressure = 75 psiGas 2 Pressure = 75 psi Curtain gas pressure = 40 psi Quadrupoleresolution low → low Transitions 1203.0 → 99.9 m/z: Cyclosporin A (CE:125 eV, CXP: 16 V) 1215.0 → 99.9 m/z: D12-Cyclosporin A (CE: 125 eV,CXP: 16 V) 1219.0 → 224.0 m/z: AM1 (CE: 65 eV, CXP: 15 V) 1219.0 → 99.9m/z: AM9 (CE: 125 eV, CXP: 16 V) 1189.0 → 224.0 m/z: AM4N (CE: 65 eV,CXP: 15 V) DP (declustering potential) 130 V

Data Evaluation

Concentrations were evaluated using an internal standard method.

The concentrations of each analyte were determined using the followingregression model, weighting factor and formula:

Weighting Analyte Regression model factor Formula for concentration all4 y = ax² + bx + c 1/conc.${concentration} = \frac{{- b} \pm \sqrt{b^{2} - {4{a\left( {c - {{peak}\mspace{14mu}{area}\mspace{14mu}{ratio}}} \right)}}}}{2a}$

Based thereon (arithmetic) mean values and relative standard deviations(CV) (formulas shown below) were calculated using the program “Lotus123”.

${{standard}\mspace{14mu}{deviation}} = \sqrt{\frac{1}{N - 1}{\sum\limits_{i = 1}^{N}\left( {x_{i} - \overset{\_}{x}} \right)^{2}}}$x_(i)  calculated  concentration$\overset{\_}{x}\mspace{31mu}{mean}\mspace{14mu}{calculated}\mspace{14mu}{concentration}$N   number  of  values i   index  of  value${{relative}\mspace{14mu}{standard}\mspace{14mu}{deviation}\mspace{14mu}(\%)} = {\frac{{{standard}\mspace{14mu}{deviation}}\;}{{mean}\mspace{14mu}{calculated}\mspace{14mu}{concentration}}*100}$

The concentration of cyclosporin A and the metabolites AM4N, AM9 infaecal samples collected on day 2 of the study is shown in Table 22.

TABLE 22 Total 2 Mean Mean Mean Total 1 CyA + CyA AM4N AM9 AM4N + AM9AM4N + AM9 Total 1/ Ratio Group (ng/g) (ng/g) (ng/g) (ng/g) (ng/g) Total2% CyA:AM4:AM9 IV Group 1 2479.58 2194.52 3343.21 5537.73 8017.31 69.1%0.45 IV Group 2 1215.75 1110.19 2942.91 4053.1 5268.85 77.9% 0.30 37.5mg OD 351258.5 1918.8 3687.9 5606.7 356865.20 1.6% 62.65 75 mg OD122940.4 1419.76 2384.75 3804.51 126744.91 3.0% 32.31 75 mg BID 159430.61069.01 2431.42 3500.43 162931.00 2.2% 45.55 150 mg BID 1068136 4388.839969.08 14357.91 1082493.61 1.3% 74.39

Pharmacokinetic Evaluation Demographic and Other BaselineCharacteristics

Demographic characteristics are summarised in Table 23. All subjectswere male and aged between 19 and 54 years. Demographic characteristicswere similar across the treatment groups; any differences were notconsidered to affect the results of the study.

TABLE 23 Demographics Sandimmun ® Sandimmun ® CyCol ® CyCol ® CyCol ®CyCol ® IV IV 37.5 mg 75 mg 75 mg 150 mg Group 1 Group 2 once daily oncedaily BID BID Overall N = 8 N = 8 N = 8 N = 8 N = 8 N = 8 N = 48 Age,years Mean 30.4 29.4 37.4 38.8 32.1 31.9 33.3 SD 9.21 7.95 10.81 12.446.22 6.92 9.38 Median 27.0 28.5 36.5 36.5 32.5 30.0 32.0 Min, Max 23, 5019, 39 21, 54 20, 54 22, 44 24, 45 19, 54 Age categories, n (%) 18-30 5(62.5) 4 (50.0) 2 (25.0) 2 (25.0) 3 (37.5) 5 (62.5) 21 (43.8) 31-55 3(37.5) 4 (50.0) 6 (75.0) 6 (75.0) 5 (62.5) 3 (37.5) 27 (56.3) Weight, kgMean 75.0 77.3 78.4 84.5 79.4 73.9 78.1 SD 9.28 12.93 15.61 13.91 11.1411.79 12.41 Median 70.8 72.3 77.4 87.1 83.7 73.7 77.4 Min, Max 65, 89 66, 103  58, 100  64, 100 59, 91 54, 91  54, 103 Height, cm Mean 176.0175.9 177.3 181.4 180.4 175.4 177.7 SD 5.76 6.75 9.33 9.47 5.07 6.057.27 Median 176.0 173.0 179.0 180.5 179.0 176.5 177.0 Min, Max 166, 185169, 188 160, 191 171, 200 174, 188 163, 183 160, 200 Race, n (%) Black0 0 1 (12.5) 1 (12.5) 2 (25.0) 1 (12.5)  5 (10.4) Caucasian 4 (50.0) 6(75.0) 4 (50.0) 5 (62.5) 6 (75.0) 6 (75.0) 31 (64.6) Asian/Pacific 3(37.5) 2 (25.0) 3 (37.5) 1 (12.5) 0 1 (12.5) 10 (20.8) Islander Mixed 1(12.5) 0 0 1 (12.5) 0 0 2 (4.2) Ethnicity, n (%) Not Hispanic  8 (100.0)7 (87.5)  8 (100.0)  8 (100.0)  8 (100.0)  8 (100.0) 47 (97.9) Hispanic0 1 (12.5) 0 0 0 0 1 (2.1) Body mass index, kg/m² Mean 24.2 24.6 24.925.6 24.4 23.9 24.6 SD 2.89 2.71 3.84 2.80 3.11 3.21 2.99 Median 23.523.5 25.8 26.1 24.1 23.2 24.1 Min, Max 21, 29 22, 29 19, 29 21, 30 20,29 20, 29 19, 30 BID = twice daily; IV = intravenous; Max = maximum; Min= minimum; n = number of subjects in assigned category; N = number ofsubjects in Safety Population; SD = standard deviation. Percentages arebased on the number of subjects in the Safety Population.

Pharmacokinetic Results and Tabulations of Individual Subject DataAnalysis of Pharmacokinetics Cyclosporin Pharmacokinetics

Median whole blood cyclosporin concentration-time profiles(linear-linear) for the CyCol® groups on Day 1 and Day 7 are provided inFIGS. 11 and 12 respectively. Cyclosporin concentrations generallyincreased with increasing dose of Cycol®.

Single Dose Pharmacokinetics

Whole blood cyclosporin pharmacokinetic parameters following a singledose are summarised in Table 24. Exposure (AUC_(inf)) was considerablylower following treatment with all doses of CyCol® compared withSandimmun® IV. Cyclosporin concentrations (AUC_(last) and AUC_(inf))increased with increasing dose of CyCol®. Median T_(max) was similar atall doses of CyCol® (between 5.0 and 5.5 hours).

Median percent extrapolation was low in the two Sandimmun® IV groups(<5%). Median percent extrapolation was highest in the CyCol® 75 mg BIDand 150 mg BID groups (24.7% and 28.9%, respectively.

TABLE 24 Summary of Single Dose Whole Blood Cyclosporin PharmacokineticParameters - PK Population Sandimmun ® Sandimmun ® CyCol ® CyCol ®CyCol ® CyCol ® IV IV 37.5 mg 75 mg 75 mg 150 mg Group 1 Group 2 oncedaily once daily BID BID N = 8 N = 8 N = 8 N = 8 N = 8 N = 8 AUC_(last)(ng · h/mL) n 8 8 8 8 Arithmetic mean 118.1 241.2 250.5 554.3 SD 62.10158.71 162.89 355.15 Geometric mean 104.7 191.7 199.1 463.7 CV % 52.665.8 65.0 64.1 Median 96.9 208.5 228.0 437.5 Minimum, Maximum 46.4, 228 55.6, 470  64.8, 480   146, 1231 AUC_(inf) (ng · h/mL) n 8 8 8 8 8 8Arithmetic mean 8836.0 8397.3 132.8 265.2 329.5 821.8 SD 1180.58 1609.7670.81 163.83 208.07 460.76 Geometric mean 8769.5 8274.5 117.9 218.3265.9 700.4 CV % 13.4 19.2 53.3 61.8 63.1 56.1 Median 8649.5 8110.5105.9 228.0 314.5 754.0 Minimum, Maximum 7457, 11088 6316, 11845 55.0,263  67.9, 502  100, 637  220, 1661 C_(max) (ng/mL) n 8 8 8 8 Arithmeticmean 18.38 41.89 28.84 59.44 SD 9.810 32.574 33.206 54.306 Geometricmean 15.70 29.86 17.66 40.93 CV % 53.4 77.8 115.2 91.4 Median 8649.58110.5 105.9 228.0 314.5 754.0 Minimum, Maximum 7457, 11088 6316, 1184555.0, 263  67.9, 502  100, 637  220, 1661 C_(max) (ng/mL) n 8 8 8 8Arithmetic mean 18.38 41.89 28.84 59.44 SD 9.810 32.574 33.206 54.306Geometric mean 15.70 29.86 17.66 40.93 CV % 53.4 77.8 115.2 91.4 Median17.70 30.40 14.40 36.60 Minimum, Maximum 6.03, 30.8 7.20, 86.9 3.73,101  10.5, 156  t_(1/2) (h) n 8 8 Arithmetic mean 5.04 3.83 SD 0.7530.556 CV % 14.9 14.5 Median 4.87 3.88 Minimum, Maximum 4.25, 6.28  2.70,4.42  T_(max) (h) n 8 8 8 8 Median 5.0 5.5 5.0 5.5 Minimum, Maximum 4, 8 4, 12  4, 12 3, 8 AUC_(0-T) (ng · h/mL) n 8 8 Arithmetic mean 8448 8122SD 1081.5 1562.3 Geometric mean 8389 8002 CV % 12.8 19.2 Median 84227867 Minimum, Maximum 7119, 10472 6054, 11465 F (%) n 8 8 8 8 Arithmeticmean 6.006 6.000 3.729 4.650 SD 3.2020 3.7087 2.3575 2.6082 Geometricmean 5.336 4.938 3.007 3.962 CV % 53.3 61.8 63.2 56.1 Median 4.790 5.1503.560 4.265 Minimum, Maximum 2.49, 11.9 1.54, 11.4 1.13, 7.21 1.24, 9.40C_(min) (ng/mL) n 8 8 8 8 Arithmetic mean 1.21 2.67 4.23 11.32 SD 0.6910.886 1.979 7.461 Geometric mean 1.04 2.53 3.73 9.69 CV % 57.2 33.2 46.965.9 Median 1.12 2.44 4.26 8.74 Minimum, Maximum 0.333, 2.58  1.37, 4.001.17, 7.40 5.06, 27.2 Percent extrapolated (%) n 8 8 8 8 8 8 Median 4.53.3 11.6 9.0 24.7 29.8 Minimum, Maximum 1.64, 6.03  2.27, 4.14  7.15,15.6 5.75, 23.7 10.6, 35.7 17.2, 56.6 BID = twice daily; IV =intravenous; N = number of subjects in Safety Population.

Steady State Pharmacokinetics

Whole blood cyclosporin pharmacokinetic parameters at steady state aresummarised in Table 25. Cyclosporin concentrations (AUC_(0-t)) increasedwith increasing dose of CyCol®. Median T_(max) was similar at all dosesof CyCol® (between 5.5 and 6.0 hours).

TABLE 25 Summary of Steady State Whole Blood Cyclosporin PharmacokineticParameters - PK Population CyCol ® CyCol ® CyCol ® CyCol ® 37.5 mg 75 mg75 mg 150 mg once daily once daily BID BID N = 8 N = 8 N = 8 N = 8AUC_(0-T) (ng · h/mL) n 8 8 7 7 Arithmetic mean 128 225 298 585 SD 53.6113.5 121.4 228.1 Geometric mean 117 190 266 539 CV % 41.9 50.4 40.739.0 Median 130 257 343 608 Minimum, Maximum 49.3, 223  54.2, 354  84.2,410  228, 886 C_(max) (ng/mL) n 8 8 7 7 Arithmetic mean 19.47 38.5736.67 45.30 SD 10.155 24.028 13.866 25.682 Geometric mean 16.63 28.8933.72 37.95 CV % 52.2 62.3 37.8 56.7 Median 21.25 48.50 41.10 48.40Minimum, Maximum 6.23, 35.9 4.94, 75.4 14.1, 49.7 12.9, 82.1 Tmax (h) n8 8 7 7 Median 5.5 6.0 6.0 6.0 Minimum, Maximum 5, 8 2, 6 1, 6 4, 8 Cmin(ng/mL) n 8 8 7 7 Arithmetic mean 1.11 2.68 5.69 10.06 SD 0.509 1.2793.355 1.859 Geometric mean 1.01 2.39 4.84 9.90 CV % 46.0 47.8 58.9 18.5Median 1.02 2.58 4.63 9.70 Minimum, Maximum 0.56, 1.99 1.17, 4.50 2.28,10.4 6.90, 12.2 F (%) n 8 8 7 7 Arithmetic mean 5.789 5.099 3.374 3.313SD 2.4267 2.5700 1.3748 1.2895 Geometric mean 5.313 4.300 3.012 3.049 CV% 41.9 50.4 40.7 38.9 Median 5.840 5.810 3.880 3.440 Minimum, Maximum2.23, 10.1 1.23, 8.01 0.95, 4.64 1.29, 5.01 C_(av) (ng/mL) n 8 8 7 7Arithmetic mean 5.33 9.38 24.86 48.77 SD 2.234 4.728 10.116 19.006Geometric mean 4.89 7.91 22.20 44.89 CV % 41.9 50.4 40.7 39.0 Median5.40 10.69 28.58 50.67 Minimum, Maximum 2.05, 9.29  2.26, 14.75  7.02,34.17 19.00, 73.83 Linearity Ratio n 8 8 7 7 Arithmetic mean 1.295 1.4030.968 0.883 SD 0.7862 1.4995 0.4929 0.3378 Geometric mean 0.996 0.8700.871 0.823 CV % 60.7 106.9 50.9 38.3 Median 1.5 0.9 0.7 0.8 Minimum,Maximum 0.238, 2.309 0.154, 4.786 0.492, 1.762 0.422, 1.300 BID = twicedaily; IV = intravenous; N = number of subjects in Safety Population.

Colon Tissue Distribution Cyclosporin Distribution

Tissue, mucous and intracolonic cyclosporin concentrations aresummarised in Table 26.

Tissue cyclosporin concentrations generally increased with increasingdose of CyCol® and concentrations were higher in the CyCol® 75 mg BIDand 150 mg BID groups than in the Sandimmun® IV groups. There was norelationship between tissue cyclosporin concentrations and distance fromanal verge.

Mucous cyclosporin concentrations were higher in the CyCol® 150 mg groupcompared with the other CyCol® groups and the Sandimmun® IV groups.There was no relationship between mucous cyclosporin concentrations anddistance from anal verge.

Intracolonic faecal cyclosporin concentrations generally increased withincreasing dose of CyCol® and were considerably higher in all of theCyCol® groups compared with the Sandimmun® IV groups.

TABLE 26 Summary of Tissue, Mucous and Intracolonic Faecal CyclosporinConcentrations - PK Population Sandimmun ® Sandimmun ® CyCol ® CyCol ®CyCol ® CyCol ® IV IV 37.5 mg 75 mg 75 mg 150 mg Group 1 Group 2 oncedaily once daily BID BID N = 8 N = 8 N = 8 N = 8 N = 8 N = 8 Tissuecyclosporin concentrations (ng/g) n 8 8 8 8 7 7 Arithmetic mean 802 11001045 1094 1579 5210 SD 397.0 276.1 712.5 881.3 502.3 4417.5 Geometricmean 717 1071 863 752 1493 4043 CV % 49.5 25.1 68.2 80.6 31.8 84.8Median 789 974 727 906 1576 3114 Minimum, Maximum 326, 1487 836, 1541352, 2431 139, 2497 669, 2279 1945, 14269 Mucous cyclosporinconcentrations (ng/g) n 8 8 8 8 7 7 Arithmetic mean 73.68 126.93 103.56101.09 78.27 596.38 SD 47.311 60.086 58.416 63.478 40.067 458.366Geometric mean 60.46 116.36 90.72 70.42 69.93 457.16 CV % 56.0 43.6 49.062.2 43.1 98.2

Table 26 shows that oral administration of the CyCol™ compositionscomprising the surfactant provided similar or higher cyclosporin Aconcentrations in colonic tissue compared to IV administration of 2mg/kg cyclosporin. However, the CyCol™ compositions resulted insignificantly lower systemic exposure compared to IV administration ofcyclosporin (see the AUC and C_(max) values in Tables 24 and 25).Reference to Table 22 also shows that the CyCol™ compositions resultedin much lower cyclosporin metabolism as evidenced by the high ratio ofcyclosporin to the AM4+AM9 metabolites in collected faecal samples.

AM1 Concentrations

Tissue, mucous and intracolonic faecal AM1 concentrations are summarisedin Table 27.

Tissue AM1 concentrations increased with increasing dose of CyCol® butwere slightly lower in the CyCol® 150 mg BID compared with theSandimmun® IV groups. There was no relationship between tissue AM1concentrations and distance from anal verge.

Mucous AM1 concentrations were similar in the CyCol® 150 mg BID andSandimmun® IV groups and lower in the other CyCol® groups. There was norelationship between mucous AM1 concentrations and distance from analverge.

Intracolonic faecal AM1 concentrations were similar in all treatmentgroups.

TABLE 27 Summary of Tissue, Mucous and Intracolonic Faecal AM1Concentrations - PK Population Sandimmun ® Sandimmun ® CyCol ® CyCol ®CyCol ® CyCol ® IV IV 37.5 mg 75 mg 75 mg 150 mg Group 1 Group 2 oncedaily once daily BID BID N = 8 N = 8 N = 8 N = 8 N = 8 N = 8 Tissue AM1concentrations (ng/g) n 8 8 8 8 7 7 Arithmetic mean 517.6 345.2 70.0101.7 232.7 303.7 SD 294.49 89.95 42.48 74.12 198.53 156.50 Geometricmean 453.4 334.5 61.8 82.5 169.3 272.5 CV % 56.9 26.1 60.7 72.9 85.351.5 Median 534.9 320.3 63.0 75.9 155.5 330.7 Minimum, Maximum   217,1143.0 205.9, 467.8 34.3, 166.3 27.8, 258.2 38.0, 624.6 132.6, 605.0Mucous AM1 concentrations (ng/g) n 8 8 8 8 7 7 Arithmetic mean 304.3307.3 60.3 67.0 56.9 233.8 SD 146.10 163.66 35.63 33.47 43.74 102.73Geometric mean 275.6 276.5 53.7 61.0 46.1 215.7 CV % 48.0 53.3 59.1 50.076.8 43.9 Median 268.1 245.1 48.6 61.2 44.3 203.3 Minimum, Maximum131.2, 566.1 169.4, 568.8 31.6, 141.1 32.9, 138.9 21.2, 145.4 122.0,378.6 Intracolonic faecal AM1 concentrations (ng/g) n 8 8 8 8 7 7Arithmetic mean 2472 4944 2939 4719 5386 5870 SD 1477.2 3375.2 645.82618.3 5528.3 3249.3 Geometric mean 2094 4029 2876 3921 3564 5236 CV %59.8 68.3 22.0 55.5 102.6 55.4 Median 2160 3322 2938 4424 4441 4933Minimum, Maximum 1027, 4712  1885, 10456 2158, 3749  1002, 8838   948,16997  3041, 12251

AM4N Concentrations

Tissue, mucous and intracolonic faecal AM4N concentrations aresummarised in Table 28.

Tissue AM4N concentrations generally increased with increasing dose ofCyCol®. Concentrations in the CyCol® 150 mg BID were lower compared withthe Sandimmun® IV groups. There was no relationship between tissue AM4Nconcentrations and distance from anal verge.

Mucous AM4N concentrations were higher in the CyCol® 150 mg BID groupcompared with the other CyCol® groups but concentrations were highest inthe Sandimmun® IV groups. There was no relationship between mucous AM4Nconcentrations and distance from anal verge. Intracolonic faecal AM4Nconcentrations were similar in all treatment groups.

TABLE 28 Summary of Tissue, Mucous and Intracolonic Faecal AM4NConcentrations - PK Population Sandimmun ® Sandimmun ® CyCol ® CyCol ®CyCol ® CyCol ® IV IV 37.5 mg 75 mg 75 mg 150 mg Group 1 Group 2 oncedaily once daily BID BID N = 8 N = 8 N = 8 N = 8 N = 8 N = 8 Tissue AM4Nconcentrations (ng/g) n 8 8 8 8 7 7 Arithmetic mean 19.816 24.272 3.0133.862 3.657 9.686 SD 23.4043 28.1267 2.8647 3.4242 2.8473 10.9795Geometric mean 11.746 11.188 2.224 2.247 2.809 6.146 CV % 118.1 115.995.1 88.7 77.9 113.4 Median 12.011 15.464 1.581 3.014 2.246 4.523Minimum, Maximum 3.49, 71.45 1.36, 81.84 1.12, 8.99 0.40, 8.41 0.78,9.11  2.30, 31.58 Mucous AM4N concentrations (ng/g) n 8 8 8 8 7 7Arithmetic mean 3.787 10.458 1.194 1.338 0.632 2.931 SD 1.7635 14.43650.9826 0.8601 0.4757 1.8756 Geometric mean 3.440 4.332 0.913 0.974 0.5132.510 CV % 46.6 138.0 82.3 64.3 75.2 64.0 Median 3.291 5.455 0.797 1.2750.385 2.179 Minimum, Maximum 1.64, 6.95  0.40, 43.78 0.34, 2.85 0.19,2.31 0.28, 1.44 1.41, 5.90 Intracolonic faecal AM4N concentrations(ng/g) n 8 8 8 8 7 7 Arithmetic mean 1390 2739 2670 2143 2134 2788 SD665.9 983.7 453.1 819.3 1554.1 1123.3 Geometric mean 1285 2599 2639 19871768 2627 CV % 47.9 35.9 17.0 38.2 72.8 40.3 Median 1119 2536 2599 22822020 2685 Minimum, Maximum 875, 2863 1785, 4554  2158, 3425  895, 3171 815, 5361 1842, 5060

AM9 Concentrations

Tissue, mucous and intracolonic faecal AM9 concentrations are summarisedin Table 29.

Tissue AM9 concentrations generally increased with increasing dose ofCyCol®. Concentrations in the CyCol® 150 mg BID were lower compared withthe Sandimmun® IV groups. There was no relationship between tissue AM4Nconcentrations and distance from anal verge.

Mucous AM9 concentrations were higher in the CyCol® 150 mg BID groupcompared with the other CyCol® groups but concentrations were highest inthe Sandimmun® IV groups. There was no relationship between mucous AM9concentrations and distance from anal verge.

Intracolonic faecal AM9 concentrations were similar in all treatmentgroups.

TABLE 29 Summary of Tissue, Mucous and Intracolonic Faecal AM9Concentrations - PK Population Sandimmun ® Sandimmun ® CyCol ® CyCol ®CyCol ® CyCol ® IV IV 37.5 mg 75 mg 75 mg 150 mg Group 1 Group 2 oncedaily once daily BID BID N = 8 N = 8 N = 8 N = 8 N = 8 N = 8 Tissue AM9concentrations (ng/g) N 8 8 8 8 7 7 Arithmetic mean 64.41 80.81 11.8115.80 28.53 50.28 SD 57.022 74.901 8.842 12.491 36.438 44.724 Geometricmean 45.98 53.22 9.57 11.38 18.15 36.42 CV % 88.5 92.7 74.9 79.1 127.788.9 Median 38.90 49.76 8.37 13.10 13.52 23.97 Minimum, Maximum  15.21,159.11 15.18, 208.30 4.49, 26.72 3.17, 37.45 5.38, 109.85 15.92, 120.30Mucous AM9 concentrations (ng/g) N 8 8 8 8 7 7 Arithmetic mean 31.48193.81 9.20 11.31 9.62 29.57 SD 16.406 180.450 6.570 7.819 9.460 18.643Geometric mean 28.40 134.27 7.70 8.88 7.37 25.14 CV % 52.1 93.1 71.469.1 98.3 63.0 Median 27.82 100.41 6.45 9.21 6.75 20.38 Minimum, Maximum15.76, 66.11 30.83, 558.92 4.23, 21.96 2.82, 25.20 3.77, 30.64  10.73,61.55  Intracolonic faecal AM9 concentrations (ng/g) N 8 8 8 8 7 7Arithmetic mean 2254 5646 3637 5479 5862 6351 SD 1121.2 4390.0 1215.23550.3 6721.1 4584.7 Geometric mean 2002 4279 3464 4292 3820 5296 CV %49.7 77.8 33.4 64.8 114.7 72.2 Median 2116 3226 3352 6123 4023 5516Minimum, Maximum 1032, 3916 1878, 11714 2158, 5396   912, 12100 953,20598 2438, 15941

Pharmacokinetic Conclusions

-   -   Systemic exposure to cyclosporin A was lower following treatment        with CyCol® at doses up to 150 mg BID once daily for 7 days        compared with a single IV infusion of Sandimmun® 2 mg/kg over 24        hours (2 mg/kg/day).    -   Tissue and mucous concentrations of cyclosporin A were higher in        the CyCol® 75 mg BID and 150 mg BID groups compared with the        Sandimmun® IV groups suggesting that these doses should be        efficacious. Tissue and mucous concentrations of AM1, AM4N and        AM9 were generally higher following treatment with Sandimmun® IV        compared with CyCol®.    -   Intracolonic faecal concentrations of cyclosporin A were        considerably higher in the CyCol® groups compared with the        Sandimmun® IV groups indicating that CyCol® is predominantly        excreted in faeces.

DISCUSSION AND OVERALL CONCLUSIONS

Systemic exposure to cyclosporin was lower following treatment withCyCol® at doses up to 150 mg BID once daily for 7 days compared with asingle IV infusion of Sandimmun® 2 mg/kg over 24 hours (2 mg/kg/day).This may result in a lower incidence of side effects related tocyclosporin following treatment with CyCol® compared with Sandimmun®.

Tissue and mucous concentrations of cyclosporin were higher in theCyCol® 75 mg BID and 150 mg BID groups compared with the Sandimmun® IVgroups suggesting that these doses should be efficacious. Tissue andmucous concentrations of AM1, AM4N and AM9 were generally higherfollowing treatment with Sandimmun® IV compared with CyCol®.

Intracolonic faecal concentrations of cyclosporin were considerablyhigher in the CyCol® groups compared with the Sandimmun® IV groupsindicating that CyCol® is predominantly excreted in faeces.

Administration of CyCol® at doses up to 150 mg BID was generally welltolerated; the majority of AEs were mild and no severe or serious AEswere reported. There was a higher incidence of gastrointestinaldisorders at the highest dose of CyCol® but none led to discontinuation.

Comparative Example 10: In-Vitro Study Using Minibead CompositionComprising Cremophor Preparation of Minibead Modified ReleaseCompositions

Minibead Formulations I and II and a Fast Release Formulation wereprepared using an analogous process to that described above.

Formulation I: “Medium” coating level (10% weight gain Opadry subcoat;11% weight gain Surelease™/Pectin overcoat)

Component % Core Cyclosporin A 8.8 Miglyol 810 N 3.8 Transcutol HP 13.5Kolliphor ™ EL 7.6 SDS 3.3 Sorbitol 4.7 Gelatin 40.3 Sub-Coat Opadry 8.2Overcoat Surelease ™ (solid contents) 9.7 Pectin 0.2Formulation II: “High” coating level (10% weight gain Opadry subcoat;17% weight-gain Surelease™/Pectin overcoat)

Component % Core Cyclosporin A 8.4 Miglyol 810 N 3.6 Transcutol HP 12.8Kolliphor ™ EL 7.2 SDS 3.1 Sorbitol 4.4 Gelatin 38.3 Sub-coat Opadry 7.8Overcoat Surelease ™ (solid contents) 14.2 Pectin 0.3

Fast Release Formulation (No Surelease™ Pectin Overcoat)

Component % Core Cyclosporin A 9.8 Miglyol 810 N 4.2 Transcutol HP 14.9Kolliphor ™ EL 8.4 SDS 3.6 Sorbitol 5.2 Gelatin 44.8 Sub-coat Opadry 9.1

Human Pharmacokinetic Study Study Objectives:

Objective 1: To compare the rate and extent of absorption ofcyclosporin-A following administration of the Fast Release Formulation(fast-release capsule; Test 1), Formulation I (medium-release capsule;Test 2), and Formulation II (slow-release capsule; Test 3) with Neoral™immediate-release capsule (reference), administered as a single 75 mgdose under fasting conditions.

Objective 2: To evaluate the amount of unchanged cyclosporin-A excretedin the faeces after administration of the Comparative Formulation(fast-release capsule; test 1), Formulation I (medium-release capsule;Test 2), Formulation II (slow-release capsule; Test 3) versus Neoral,administered as a single 75 mg dose under fasting conditions.

Study Design:

A single centre, randomised, single-dose, open-label, 4-period,4-sequence crossover comparative BA study, performed under fastingconditions.

Subjects:

Enrolled and randomised: 18 (12 females and 6 males) Withdrew consent: 0

Withdrawal: 1 (was withdrawn) Completed all 4 periods: 16

Safety population: 18

Pharmacokinetic (PK) population: 18

Pharmacokinetic Analysis

The mean pharmacokinetic values obtained in the study are summarized inthe Table 30.

TABLE 30 Summary of pharmacokinetic parameters for cyclosporin-A foreach treatment Mean ± SD (CV %) Whole Blood Cyclosporin-A Cyclosporin-A(Test 1) Cyclosporin-A Cyclosporin-A Fast Release (Test 2) (Test 3)Formulation Formulation I Formulation II Neoral N 17   17   18   17  AUC_(0-t) 1212.52 ± 297.62 609.89 ± 280.15 408.49 ± 231.01 1582.20 ±358.09  (ng · hr/mL) (24.55) (45.93) (56.55) (22.63) AUC_(0-inf) 1257.83± 312.14 672.07 ± 296.71 474.37 ± 247.93 1639.78 ± 371.52  (ng · hr/mL)(24.82) (44.15) (52.27) (22.66) C_(max) 321.33 ± 87.61 138.28 ± 63.54 82.81 ± 48.01 594.66 ± 117.01 (ng/mL) (27.27) (45.95) (57.98) (19.68)Residual Area  3.55 ± 0.71 10.72 ± 8.10  15.38 ± 12.69 3.52 ± 0.77 (%)(20.12) (75.50) (82.52) (21.87) T_(max) ^(a)  2.00  5.00  5.00  1.25(hr) (1.25-3.00) (5.00-8.00) (5.00-10.0) (1.00-1.75) K_(el)  0.1105 ±0.0113 0.0863 ± 0.0259 0.0822 ± 0.0232 0.1037 ± 0.0103 (1/hr) (10.25)(30.01) (28.20)  (9.97) T_(1/2 el)  6.33 ± 0.61 8.72 ± 2.76 9.49 ± 4.556.75 ± 0.77 (hr)  (9.70) (31.66) (47.96) (11.43) ^(a)Median (Min-Max) InTable 30 the AUC and Cmax values are the mean value ± standard deviation(SD)

The whole blood concentration of cyclosporin A for each of thesecomposition is shown in FIGS. 13A-B.

A comparison of FIGS. 11 and 12 using the minibeads of the presentinvention containing Capmul Capmul GMO-50 (glyceryl monooleate/dioleate)as the surfactant with FIGS. 13A-13B (and the corresponding data tables)shows that the Capmul containing composition according to the inventionprovided lower C_(max) and AUC values indicating lower systemic exposureto cyclosporin A.

Determination of Cyclosporin-A and its Metabolites, AM9 and AM4N, inFaecal Samples

Faecal samples collected during the PK trial were analysed byRP-LC-MS/MS as described previously. The results are shown in Table 31.

TABLE 31 Total 2 Mean Mean Mean Total 1 CyA + Total 1/ Ratio CSA % AM4N% AM9 % AM4N + AM9 % AM4N + AM9 % Total 2% CyA:AM4:AM9 Fast Release 73.812 14.2 26.2 100 26.20% 2.82 Formulation I 86.9 5.5 7.6 13.1 100 13.10%6.63 Formulation II 91.5 3.4 5.1 8.5 100 8.50% 10.76 Neoral 37.1 26.236.7 62.9 100 62.90% 0.59

A comparison of the faecal data in Table 31 with the faecal dataobtained with the Capmul containing compositions of Example 9 (Table 22)shows that the Capmul compositions of Example 9 resulted in much lowercyclosporin metabolism than the comparative compositions containingCremophor. This is illustrated by the lower relative % of the (AM9+AM4N)metabolites in the collected faeces for the Capmul compositions in Table22 compared to the Cremophor compositions in Table 31. This differenceis clearly illustrated in FIG. 14 which shows the ratio of cyclosporinto (AM4N+AM9) measured in the faecal samples for each of the testedformulations.

The compositions of the invention comprising Capmul GMO-50 resulted insignificantly less cyclosporin A metabolism compared to the compositionscontaining Cremophor. The compositions of the invention thereforeprovide higher local levels of cyclosporin in the colon as a result ofthe reduced systemic and non-systemic metabolism of the cyclosporinreleased from the composition. The compositions of the invention mayenable a lower dose of cyclosporin to be administered whilst maintaininga therapeutic effect, thereby further increasing the therapeutic window.

FIG. 15 compares the in-vitro dissolution profile of the Capmulformulation used in Example 9 with the cremophor compositions used inComparative Example 10 using the two-stage dissolution test describedherein. FIG. 15 shows that the release profiles for the Capmulcomposition, the Slow Release, and the Medium Release are all verysimilar in the 2 to 5 hour period. During this time the compositions areexpected to release cyclosporin in the small intestine and would beprone to both systemic and enteric P450 metabolism of the cyclosporin.Despite the similarities in the in-vitro release profiles, FIG. 14 showsthat the Capmul composition of the present invention significantlyreduced the metabolism of cyclosporin A compared to the Fast, Medium andSlow compositions containing Cremophor as a surfactant. Additionally theCapmul composition exhibited a lower AUC and Cmax illustrating a lowersystemic exposure to cyclosporin than the Cremophor compositionsfollowing oral administration (see Tables 24 and 30).

Example 11: Emulsion Stability and Bead Formation: Effect of Surfactantand Surfactant Concentration

Compositions with differing aqueous phase surfactants were investigated.The different aqueous phase surfactants were compared to SDS. Threedifferent families of surfactant were chosen to test: Sucrose Fatty AcidEsters, Sodium n-Alkyl Sulfates and Fatty Alcohol Ethoxylates (Brijseries).

Emulsions were prepared by mixing an oil phase and an aqueous phase, asdescribed in Example 1. The oil phase was consistent for all emulsionsof this example. The emulsion aqueous phase mixed was one of threeaqueous mixtures. The three aqueous phases differed in the amount ofsurfactant, 0.7%, 1.3% and 2.5%, and water. The oil and aqueous phasemixtures are shown in tables 32 to 34.

TABLE 32 Aqueous Phase Oil Phase Component % Component % H₂O 79.56%Transcutol HP 37.00% Gelatin 17.14% Capmul GMO 50 26.00% D-Sorbitol2.00% CyA 24.50% Surfactant 1.30% Miglyol 12.50%

TABLE 33 Aqueous Phase Oil Phase Component % Component % H₂O 79.62%Transcutol HP 37.00% Gelatin 17.14% Capmul GMO 50 26.00% D-Sorbitol2.00% CyA 24.50% Surfactant 0.70% Miglyol 12.50%

TABLE 34 Aqueous Phase Oil Phase Component % Component % H₂O 78.36%Transcutol HP 37.00% Gelatin 17.14% Capmul GMO 50 26.00% D-Sorbitol2.00% CyA 24.50% Surfactant 2.50% Miglyol 12.50%

The emulsion was prepared by mixing the oil phase and aqueous phase in aoil:aqueous phase ratio of 1:5. The experiments were performed intriplicate, N=3. In order to replicate current manufacturing conditionsas much as possible the following procedure was carried out:

-   -   Identical size beakers were used to ensure minimal variation        between experiments    -   Identical submersible magnetic stirrers were placed in each        beaker to keep the emulsions under constant magnetic stirring,        and the same rpm stir rate used for each    -   Hot plates were used to maintain a constant emulsion temperature        of 65° C.    -   Emulsions were kept covered with aluminium foil during the        process

Sampling of the emulsion was carried out at various time points usingdisposable pipettes to avoid cross contamination.

Emulsion Analysis

Samples were withdrawn from each emulsion at half hour time intervalsstarting at time zero (t0) using a disposable pipette. Drops of emulsionwere placed on a glass slide, pre heated on a hot plate. The sample wascovered with a cover slip and a small amount of pressure applied to forma thin film of emulsion. The samples were allowed to solidify to roomtemperature before being viewed at ×50 and ×100 objective lens underpolarised light to check for the presence of crystals. Results wererecorded as optical photomicrographs with three images taken of eachemulsion at each time point. Images documenting crystal growth and sizewith respect to time within the sample emulsion as well as backgroundoil droplet size were collected. Table 35, shown below, gives the timepoint when crystallisation was observed.

Photomicrographs of the emulsions enabled the investigation of thedroplet size as well as crystal formation. Smaller, more uniform dropletsize was preferred. These preferred droplet characteristics wereobserved in some of the tested emulsions, particularly the emulsionscontaining Sodium n-Octyl Sulfate and Sodium n-Octadecyl Sulfate.

The photomicrographs of an emulsion for each surfactant at specifiedtime points are shown in FIGS. 16A to 22I.

Bead Formation

Bead formation was attempted, to examine how bead formation was affectedby the different surfactants. Samples (1 ml) were withdrawn from eachemulsion using a Gilson pipette when the emulsion stability experimenthad been terminated, before the emulsions were discarded. Beads wereformed by dropping the emulsion at a steady rate into a cooling bath ofmedium chain triglyceride oil which was kept refrigerated. Beads wereretrieved using a sieve and placed on tissue paper within a container,gently patted with tissue paper to remove excess surface oil and left todry overnight on the worktop at room temperature. Bead formation wasdeemed to have occurred if a spherical or nearly spherical bead wasformed by dropping an emulsion into a cooling oil or expelling theemulsion when under the surface of the cooling oil. Formation of oval orelongated beads was not deemed as bead formation. It has to beacknowledged that due to the manual nature of the method of producingbeads employed here compared with bead production using the Spherexequipment at a manufacturing scale, formulations which may not formbeads at this lab scale study may in fact form beads when processedusing the Spherex equipment.

TABLE 35 Concen- tration in aqueous Beads Emulsion phase % For- imagesin Surfactant w/w Onset Crystallisation mation FIG. sodium 1.4 (t = 2hours Y dodecyl sulphate Sucrose 0.7 (t = 2 hours) Y 16A-16C Laurate 1.3(t = 1 hour 30 mins) N 16D-16F 2.5 (t = 1 hour) N 16G-16I Sucrose 0.7 (t= 2 hours 30 mins) Y 17A-17C Palmitate 1.3 (t = 1 hour 30 mins) Y17D-17F 2.5 (t = 30 mins) N 17G-17I Sodium 0.7 (t = 2 hours 30 mins) Y18A-18C n-Octyl 1.3 (t = 4 hours) Y 18D-18F Sulphate 2.5 (t = 1 hour 30mins) Y 18G-18I Sodium 0.7 (t = 3 hours) Y 19A-19C n-Octadecyl 1.3 (t =2 hours) Y 19D-19F Sulfate 2.5 (t = 2 hours) Y 19G-19I Brij L4 0.7 (t =1 hour 30 mins) Y 20A-20C (Polyethylene 1.3 (t = 2 hours 30 mins) N20D-20F Glycol 2.5 (t = 1 hour 30 mins) N 20G-20I Dodecyl Ether) BrijC10 0.7 (t = 1 hour 30 mins) N 21A-21C (PEG 1.3 (t = 1 hour) N 21D-21FHexadecyl 2.5 (t = 1 hour) N 21G-21I Ether) Brij S10 0.7 (t = 1 hour) Y22A-22C (PEG 1.3 (t = 1 hour) N 22D-22F Octadecyl 2.5 (t = 1 hour) N22G-22I Ether)

The anionic surfactants Sodium n-Octyl Sulfate and Sodium n-OctadecylSulfate in particular provided stability that was comparable to that ofsodium dodecyl sulfate. Sodium n-Octadecyl Sulfate provided the beststability for each concentration. Non-ionic surfactants providedemulsion stability but with more rapid onset of crystallization comparedto SDS, except for Brij L4 at 1.3% aqueous concentration which gave anincrease in emulsion stability.

1-119. (canceled)
 120. A method of treating a condition of thegastrointestinal tract in a subject, wherein the method comprisesadministering to the subject a composition comprising cyclosporin, ahydrogel forming polymer matrix, a surfactant and an oil phase beingdispersed in the hydrogel forming polymer matrix, wherein the surfactantcomprises a medium chain or long chain fatty acid mono- or di-glycerideor a combination thereof and does not comprise a polyethyleneglycolether or ester; wherein the hydrogel-forming polymer is in an amount ofat least 25% by weight based upon the dry weight of the composition.121. The method of claim 120, wherein the condition of thegastrointestinal tract is selected from: inflammatory bowel disease,irritable bowel disease, Crohn's disease, ulcerative colitis, celiacdisease, graft vs host disease, gastrointestinal graft-versus-hostdisease, gastroenteritis, duodenitis, jejunitis, ileitis, peptic ulcer,Curling's ulcer, appendicitis, colitis, pseudomembraneous colitis,diverticulosis, diverticulitis, collagenous colitis, endometriosis,colorectal carcinoma and adenocarcinoma.
 122. The method of claim 121,wherein the condition of the gastrointestinal tract is ulcerativecolitis.
 123. The method of claim 120, wherein the oil phase comprises asolution of the cyclosporin.
 124. The method of claim 120, wherein thesurfactant is present in an amount of more than 6 wt % of the dry weightof the composition.
 125. The method of claim 120, wherein the surfactantis selected from: glyceryl monocaprate, glyceryl dicaprate, glycerylmonocaprylate, glyceryl dicaprylate, glyceryl caprate, glycerylmonocaprylate/caprate, glyceryl caprylate/caprate glyceryldicaprylate/caprate, glyceryl monooleate/dioleate, glyceryl monooleate,glyceryl dioleate, glyceryl monostearate, glyceryl distearate, glycerylmonopalmitostearate, glyceryl dipalmitostearate, glyceryl monobehenate,glyceryl dibehenate, glyceryl monolinoleate, glycerol monolinoleate,glyceryl dilinoleate, polyglyceryl dioleate, propylene glycolmonoheptanoate, polyglycerol dioleate, or a combination thereof. 126.The method of claim 120, wherein the surfactant is selected from:glyceryl monocaprylate/caprate, glyceryl dicaprylate/caprate, glycerylmonooleate, glycerol monolinoleate, glyceryl dioleate, glycerylmonostearate, glyceryl distearate, glyceryl monopalmitostearate,glyceryl dipalmitostearate, glyceryl monobehenate, glyceryl dibehenate,glyceryl monolinoleate, glyceryl dilinoleate, polyglyceryl dioleate or acombination thereof.
 127. The method of claim 120, wherein thesurfactant is selected from: glyceryl caprylate/caprate (Capmul MCM),glyceryl monooleate/dioleate (Capmul GMO-50), glycerol monolinoleate(Maisine 35-1) or a combination thereof.
 128. The method of claim 120,wherein the surfactant is glyceryl caprylate, glyceryl caprate, glycerylmonooleate, glyceryl dioleate, glycerol monolinoleate or a combinationthereof.
 129. The method of claim 120, wherein the oil phase comprises aliquid lipid, wherein the liquid lipid comprises a medium chain fattyacid triglyceride or a combination thereof, or the liquid lipidcomprises a caprylic/capric triglyceride composition.
 130. The method ofclaim 120, wherein the composition further comprises a solvent, whereinthe solvent is miscible with the oil phase and water.
 131. The method ofclaim 120, wherein the composition further comprises a high HLBsurfactant having an HLB value of at least
 10. 132. The method of claim130, wherein the high HLB surfactant is selected from a fatty acid saltor a bile salt.
 133. The method of claim 120, wherein the hydrogelforming polymer or hydrogel forming polymer matrix comprises areversible hydrocolloid.
 134. The method of claim 120, wherein thehydrogel forming polymer or hydrogel forming polymer matrix comprises ahydrocolloid selected from carrageenan, gelatin, agar and pectin, or acombination thereof.
 135. The method of claim 120, wherein thecomposition is a solid composition and has the characteristics of acomposition formed by mixing an aqueous phase premix with an oil phasepremix to form a mixture, the aqueous phase premix comprising thehydrogel forming polymer matrix and the oil phase premix comprising theoil phase in which the cyclosporin is dissolved and the surfactant. 136.The method of claim 135, wherein the composition further comprises asurfactant having an HLB value of at least 10 which is present in theaqueous phase premix.
 137. The method of claim 120, wherein thecomposition is a solid composition and further comprises at least onecoating.
 138. The method of claim 137, wherein the at least one coatingis adapted to release the cyclosporin in at least the colon.
 139. Themethod of claim 138, wherein the coating comprises ethylcellulose. 140.The method of claim 137, wherein the at least one coating is present inan amount corresponding to a weight gain due to the coating of from 2%to 40%.
 141. The method of claim 137, wherein the composition comprisestwo coatings, a first coating and a second coating.
 142. The method ofclaim 141, wherein the first coating comprises a water-soluble celluloseether and the second coating comprises ethylcellulose.
 143. The methodof claim 142, wherein the water-soluble cellulose ether ishydroxypropylmethyl cellulose.
 144. The method of claim 120, wherein thecomposition is a solid composition that is in the form of a minibeadhaving a size of from 0.5 mm to 5 mm.
 145. The method of claim 137,wherein the composition provides a mean whole blood cyclosporin AAUC0-inf of from about 140 to about 420 ng·hr/ml.
 146. The method ofclaim 137, wherein the composition provides a Cmax of cyclosporin A offrom about 15 to about 60 ng/ml.
 147. The method of claim 137, whereinthe time taken to reach maximum whole blood concentration of cyclosporinA following oral administration of a single dose of the composition(Tmax) is between about about 3 hours to about 10 hours.
 148. The methodof claim 137, wherein the cyclosporin A absolute bioavailabilityfollowing oral administration of the composition is less than 15%. 149.The method of claim 137, wherein the composition releases less than 15%of the cyclosporin A after 2 hours; releases 10% to 40% of thecyclosporin A at 4 hours; and releases from about 30% to 70% of thecyclosporin A between 4 hours and 12 hours, when measured in a two stagedissolution test using a USP Apparatus II with a paddle speed of 75 rpmand a dissolution medium temperature of 37° C.; wherein for the first 2hours of the dissolution test the dissolution medium is 750 ml of 0.1 NHCl, and at 2 hours 250 ml of 0.2M tribasic sodium phosphate containing2% SDS is added to the dissolution medium and the pH is adjusted to pH6.8.
 150. The method of claim 137, wherein the composition provides aconcentration of cyclosporin A in colonic tissue of at least 250 ng/gfollowing oral administration of the composition to a human.
 151. Themethod of claim 150, wherein the total daily dose of cyclosporin Aadministered to the human is in the range of from 1 mg to 500 mg. 152.The method of claim 120, wherein the composition further comprises aP450 inhibitor or a PgP inhibitor, or a combination thereof.